Kinase inhibitors

ABSTRACT

A compound of the general formula: 
                         
or pharmaceutically acceptable salts, hydrates, solvates, crystal forms or diastereomers thereof is described. A method of treating protein kinase-associated disease states using the compound of formula I is also described.

CROSS-REFERENCE TO RELATED APPLICATION

This application is a divisional of U.S. application Ser. No. 10/472,156filed Sep. 19, 2003, which is the national phase of PCT applicationPCT/AU03/00628 having an international filing date of May 23, 2003,which claims benefit of U.S. provisional application No. 60/398,998filed Jul. 26, 2002 and Australian application No. PS2514 filed May 23,2002. Each of these documents is incorporated herein by reference intheir entirety.

FIELD OF THE INVENTION

The present invention relates to the field of inhibitors of proteinkinases.

BACKGROUND OF THE INVENTION

Protein kinases are a family of enzymes that catalyse thephosphorylation of specific residues in proteins. In general proteinkinases fall into several groups; those which preferentiallyphosphorylate serine and/or threonine residues, those whichpreferentially phosphorylate tyrosine residues and those whichphosphorylate both tyrosine and Ser/Thr residues. Protein kinases aretherefore key elements in signal transduction pathways responsible fortransducing extracellular signals, including the action of cytokines ontheir receptors, to the nuclei, triggering various biological events.The many roles of protein kinases in normal cell physiology include cellcycle control and cell growth, differentiation, apoptosis, cell mobilityand mitogenesis.

Protein kinases include, for example, but are not limited to, members ofthe Protein Tyrosine Kinase family (PTKs), which in turn can be dividedinto the cytoplasmic PTKs and the receptor PTKs (RTKs). The cytoplasmicPTKS include the SRC family, (including: BLK; FGR; FYN; HCK; LCK; LYN;SRC; YES and YRK); the BRK Family (including: BRK; FRK, SAD; and SRM);the CSK family (including: CSK and CTK); the BTK family, (including BTK;ITK; TEC; MKK2 and TXK), the Janus kinase family, (including: JAKI,JAK2, JAK3 and Tyk2), the FAK family (including, FAK and PYK2); the Fesfamily (including FES and FER), the ZAP70 family (including ZAP70 andSYK); the ACK family (including ACK1 and ACK2); and the Abl family(including ABL and ARG). The RTK family includes the EGF-Receptor family(including, EGFR, HER2, HER3 and HER4); the Insulin Receptor family(including INS-R and IGF1-R); the PDGF-Receptor family (includingPDGFRα, PDGFRβ, CSF1R, KIT, FLK2); the VEGF-Receptor family (including;FLT1, FLK1 and FLT4); the FGF-Receptor family (including FGFR1, FGFR2,FGFR3 and FGFR4); the CCK4 family (including CCK4); the MET family(including MET and RON); the TRK family (including TRKA, TRKB, andTRKC); the AXL family (including AXL, MER, and SKY); the TIE/TEK family(including TIE and TIE2/TEK); the EPH family (including EPHA1, EPHA2,EPHA3, EPHA4, EPHA5, EPHA6, EPHA7, EPHA8, EPHB1, EPHB2, EPHB3, EPHB4,EPHB5, EPHB6); the RYK family (including RYK); the MCK family (includingMCK and TYR010); the ROS family (including ROS); the RET family(including RET); the LTK family (including LTK and ALK); the ROR family(including ROR1 and ROR2); The Musk family (including Musk); the LMRfamily including LMR1, LMR2 and LMR3); and the SuRTK106 family(including SuRTK106).

Similarly, the serine/threonine specific kinases comprise a number ofdistinct sub-families, including; the extracellular signal regulatedkinases, (p42/ERK2 and p44/ERKI); c-Jun NH2-terminal kinase (JNK);cAMP-responsive element-binding protein kinases (CREBK); cAMP-dependentkinase (CAPK); mitogen-activated protein kinase-activated protein kinase(MAPK and its relatives); stress-activated protein kinase p38/SAPK2;mitogen-and stress-activated kinase (MSK); protein kinases, PKA, PKB andPKC inter alia.

Additionally, the genomes of a number of pathogenic organisms possessgenes encoding protein kinases. For example, the malarial parasitePlasmodium falciparum and viruses such as HPV and Hepatitis virusesappear to bear kinase related genes.

Inappropriately high protein kinase activity has been implicated in manydiseases resulting from abnormal cellular function. This might ariseeither directly or indirectly, for example by failure of the propercontrol mechanisms for the kinase, related for example to mutation,over-expression or inappropriate activation of the enzyme; or by over-or under-production of cytokines or growth factors also participating inthe transduction of signals upstream or downstream off the kinase. Inall se instances, selective inhibition of the action of the kinase mightbe expected to have a beneficial effect. Diseases where aberrant kinaseactivity has been implicated include: diabetes; restenosis;atherosclerosis; fibrosis of the liver and kidney; ocular diseases;myelo- and lymphoproliferative disorders; cancer-such as prostatecancer, colon cancer, breast cancer, head and neck cancer, leukemia andlymphoma; and, auto-immune diseases such as Atopic Dermatitis, Asthma,rheumatoid arthritis, Crohn's disease, psoriasis, Crouzon syndrome,achondroplasia, and thanatophoric dysplasia.

The JAK family of protein tyrosine kinases (PTKs) play a central role inthe cytokine dependent regulation of the proliferation and end functionof several important cell types of the immune system.

A direct comparison of the four currently known mammalian JAK familymembers reveals the presence of seven highly conserved domains (Harpuret al. 1992). In seeking a nomenclature for the highly conserved domainscharacteristic of this family of PTKs, the classification used wasguided by the approach of Pawson and co-workers (Sadovski et al, 1986)in their treatment of the SRC homology (SH) domains. The domains havebeen enumerated accordingly with most C-terminal homology domaindesignated JAK Homology domain 1 (JH1). The next domain N-terminal toJH1 is the kinase-related domain, designated here as the JH2 domain.Each domain is then enumerated up to the JH7 located at the N-terminus.The high degree of conservation of these JAK homology (JH) domainssuggests that they are each likely to play an important role in thecellular processes in which these proteins operate. However, theboundaries of the JAK homology domains are arbitrary, and may or may notdefine functional domains. Nonetheless, their delineation is a usefuldevice to aid the consideration of the overall structural similarity ofthis class of proteins.

The feature most characteristic of the JAK family of PTKs is thepossession of two kinase-related domains (JH1 and JH2) (Wilks et al,1991). The putative PTK domain of JAK1 (JH1) contains highly conservedmotifs typical of PTK domains, including the presence of a tyrosineresidue at position 1022 located 11 residues C-terminal to sub-domainVII that is considered diagnostic of membership of the tyrosine-specificclass of protein kinases. Alignment of the human JAK1 PTK domain (255amino adds), with other members of the PTK class of proteins revealedhomology with other functional PTKs (for example, 28% identity withc-fes (Wilks and Kurban, 1988) and 37% homology to TRK (Kozma et al,1988). The JH1 domains of each of the JAK family members possess ainteresting idiosyncrasy within the highly conserved sub-domain VIIImotif (residues 1015 to 1027 in JAK2) that is believed to lie close tothe active site, and define substrate specificity. The phenylalanine andtyrosine residues flanking the conserved tryptophan in this motif areunique to the JAK family of PTKs. Aside from this element, the JH1domains of each of the members of the JAK family are typical PTKdomains.

The central role played by the JAK family of protein tyrosine kinases inthe cytokine dependent regulation of the proliferation and end functionof several important cell types means that agents which inhibit JAK areuseful in the prevention and chemotherapy of disease states dependent onthese enzymes. Potent and specific inhibitors of each of the currentlyknown four JAK family members will provide a means of inhibiting theaction of those cytokines that drive immune pathologies, such as asthma(e.g. IL-13; JAK1, JAK2), and leukemia/lymphoma (e.g. IL-2: JAK1 andJAK3).

Furthermore, certain types of cancer such as prostate cancer developautocrine production of certain cytokines as a selectable mechanism ofdeveloping growth and/or metastatic potential. An example of this iscancer of the prostate, where IL-6 is produced by and stimulates thegrowth of prostate cancer cell lines such as TSU and TC3 (Spiotto MT,and Chung TD, 2000). Interestingly, levels of IL-6 are elevated in seraof patients with metastatic prostate cancer.

A great deal of literature covets the area of cytokine signalling. Thepresent inventors have focussed on the JAK/STAT pathway that is involvedin the direct connection of cytokine receptor to target genes (such ascell cycle regulators (e.g. p21) and anti-apoptosis genes (such as(Bcl-X_(L))).

The JAK/STAT Pathway

The delineation of a particularly elegant signal transduction pathwaydownstream of the non-protein tyrosine kinase cytokine receptors hasrecently been achieved. In this pathway the key components are: (i) Acytokine receptor chain (or chains) such as the Interleukin-4 receptoror the Interferon γ receptor; (ii) a member (or members) of the JAKfamily of PTKs; (iii) a member(s) of the STAT family of transcriptionfactors, and (iv) a sequence specific DNA element to which the activatedSTAT will bind.

A review of the JAK/STAT literature offers strong support to the notionthat this pathway is important for the recruitment and marshalling ofthe host immune response to environmental insults, such as viral andbacterial infection. This is well exemplified in Table 1 and Table 2.Information accumulated from gene knock-out experiments have underlinedthe importance of members of the JAK family to the intracellularsignalling triggered by a number of important immune regulatorycytokines. The therapeutic possibilities stemming from inhibiting (orenhancing) the JAK/STAT pathway are thus largely in the sphere of immunemodulation, and as such are likely to be promising drugs for thetreatment of a range of pathologies in this area. In addition to thediseases listed in Tables 1 and 2, inhibitors of JAKs could be used asimmunosuppresive agents for organ transplants and autoimmune diseasessuch as lupus, multiple sclerosis, rheumatoid arthritis, Type Idiabetes, autoimmune thyroid disorders, Alzheimer's disease and otherautoimmune diseases. Additionally, treatment of cancers such asprostate-cancer by JAK inhibitors is indicated.

TABLE 1 Cell Types Disease Type Involved Characteristics Atopy AllergicAsthma (Mast Cells T-cell activation of Atopic Dermatitis (Eczema)(Eosinophils B-cells followed by IgE Allergic Rhinitis (T-Cells mediatedactivation of (B-Cells resident Mast cells and Eosinophils Cell MediatedHypersensitivity Allergic Contact Dermatitis (T-cells HypersensitivityPneumonitis (B-cells T-cell hypersensitivity Rheumatic Diseases SystemicLupus (Monocytes Cytokine Production Erythematosus (SLE) (e.g.TNF, IL-1,CSF-1, Rheumatoid Arthritis (Macrophages GM-CSF) Juvenile Arthritis(Neutrophils T-cell Activation Sjögren's Syndrome (Mast Cells JAK/STATactivation Scleroderma (Eosinophils Polymyositis (T-Cells AnkylosingSpondylitis (B-Cells Psoriatic Arthritis Viral Diseases Epstein BarrVirus (EBV) Lymphocytes JAK/STAT Activation Hepatitis B HepatocytesJAK/STAT Activation Hepatitis C Hepatocytes JAK/STAT Inhibition HIVLymphocytes JAK/STAT Activation HTLV 1 Lymphocytes JAK/STAT ActivationVaricella-Zoster Virus (VZV) Fibroblasts JAK/STAT Inhibition HumanPapilloma Virus Epithelial cells JAK/STAT Inhibition (HPV) CancerLeukemia Leucocytes (Cytokine production Lymphoma Lymphocytes (JAK/STATActivation

TABLE 2 Diseases Potentially Treatable By JAK-Based Drug Therapies JAKfamily Strength of Target Disease Cytokine member Association AsthmaIL-4 & IL-9 JAK1 &JAK3 +++ IL-13 JAK1 & JAK2 +++ IL-5 JAK2 +++ EczemaIL-4 JAK1 & JAK3 +++ IFN-α JAK1 & JAK2 +++ Food Allergy IL-4 JAK1 & JAK3+++ Inflammatory Bowel IL-4 JAK1 & JAK3 +++ Disease & Crohn's DiseaseLeukaemia And (IL-2) JAK3, JAK1 & +++ Lymphoma JAK2 Cutaneous GM-CSF &JAK1 & JAK2 +++ Inflammation IL-6 Immune Suppression IL-10 JAK1 & TYK2+++ By Solid Tumour Prostate Cancer IL-6 JAK1, JAK2 +++ &Tyk2

BRIEF DESCRIPTION OF THE FIGURES

FIGS. 1 and 2 show the inhibitory activity of representative pyrazinesin Tel-Jak2 and Tel-Jak3 transformed cell lines, respectively. FIG. 3shows the inhibitory activity of 2-(α-methyl benzylamino)-pyrazine inTel-Jak2 transformed cell line.

SUMMARY OF THE INVENTION

The present inventors have found that a group of compounds based uponthe disubstituted pyrazine scaffold I, are inhibitors of proteinkinases.

Accordingly, in a first aspect the present invention consists in acompound of the general formula:

or pharmaceutically acceptable salts, hydrates, solvates, crystal formsor diastereomers thereof, wherein:

D is a heterocyclic ring selected from:

-   -   where X₁, X₂, X₃, X₄ are optionally substituted carbon, or one        of X₁, X₂, X₃, X₄ is N; R2 is 0-4 substituents independently        chosen from H, halogen, C₁₋₄ alkyl, CH₂F, CHF₂, CF₃, OCF₃, aryl,        hetaryl, C₁₋₄alkylOC₁₋₄ alkyl, C₁₋₄alkylOaryl, C₁₋₄alkylNR3R4,        CO₂R3, CONR3R4, CONR3SO₂R4, NR3R4, C₁₋₄, alkylNR3R4, nitro;        NR3COR4, NR5CONR3R4, NR3SO₂R4, C₁₋₄alkylNR3COR4,        C₁₋₄alkylNR5CONR3R4, C₁₋₄alkylNR3SO₂R4; and R3, R4 are each        independently H, halogen, CH₂F, CHF₂, CF₃, C₁₋₄, alkyl,        C₁₋₄alkyl cycloalkyl, C₁₋₄ cyclohetalkyl, aryl, C₁₋₄ alkyl aryl,        hetaryl, C₁₋₄ alkyl hetaryl, or may be joined to form an        optionally substituted 3-8 membered (saturated or unsaturated)        ring optionally containing an atom selected from O, S, NR6; and        R5 is selected from H, C₁₋₄alkyl, halogen, CH₂F, CHF₂, CF₃, aryl        or hetaryl; and R6 is selected from H, C₁₋₄, alkyl, aryl,        hetaryl, C₁₋₄ alkyl aryl, C₁₋₄ alkyl hetaryl.

R1 is H, C₁₋₄ alkyl, C₁₋₆ cycloalkyl.

Q is a bond, CH₂, C₁₋₄ alkyl

A is aryl, hetaryl optionally substituted with 0-3 substituentsindependently chosen from halogen, C₁₋₄ alkyl, CH₂F, CHF₂, CF₃, OCF₃,CN, NR8R9, aryl, hetaryl, C₁₋₄aryl, C₁₋₄hetaryl, C₁₋₄ alkylNR8R9, OC₁₋₄alkylNR8R9, nitro, NR10C₁₋₄NR8R9, NR8COR9, NR10CONR8R9, NR8SO₂R9,CONR8R9, CO₂R8 where R8 and R9 are each independently H, C₁₋₄ alkyl,aryl or which together form an optionally substituted 4-8 membered ringwhich may contain a heteroatom selected from O, S, NR11, where R11 isC₁₋₄ alkyl, and R10 is selected from H, C₁₋₄ alkyl.

W is selected from H, C₁₋₄alkyl, C₂₋₆alkenyl; where C₁₋₄alkyl orC₂₋₆alkenyl may be optionally substituted with C₁₋₄alkyl, OH,OC₁₋₄alkyl, NR12R13; and R12, and R13 are each independently H,C₁₋₄alkyl, or may be joined to form an optionally substituted 3-8membered ring optionally containing an atom selected from O, S, NR14 andR14 is selected from H, C₁₋₄ alkyl.

In a second aspect the present invention consists in a compositioncomprising a carrier and at least one compound of the first aspect ofthe invention.

In a third aspect the present invention consists in a method of treatinga protein kinase-associated disease state, the method comprisingadministering a therapeutically effective amount of at least onecompound of the first aspect of the invention or a therapeuticallyeffective amount of a composition of the second aspect of the invention.

In further aspects the present invention provides the use of thecompounds described in the preparation of medicaments for the treatmentof protein kinase-associated disease states.

DETAILED DESCRIPTION

In a first aspect the present invention consists in a compound of thegeneral formula:

or pharmaceutically acceptable salts, hydrates; solvates; crystal formsor diastereomers thereof, wherein:

D is a heterocyclic ring selected from:

-   -   where X₁, X₂, X₃, X₄ are optionally substituted carbon, or one        of X₁, X₂, X₃, X₄ is N; R2 is 0-4 substituents independently        chosen from H, halogen, C₁₋₄ alkyl, CH₂F, CHF₂, CF₃, OCF₃, aryl,        hetaryl, C₁₋₄alkylOC₁₋₄ alkyl, C₁₋₄alkylOaryl, C₁₋₄alkylNR3R4,        CO₂R3, CONR3R4, CONR3SO₂R4, NR3R4, C₁₋₄alkylNR3R4, nitro,        NR3COR4, NR5CONR3R4, NR3SO₂R4, C₁₋₄alkylNR3COR4,        C₁₋₄alkylNR5CONR3R4, C₁₋₄alkylNR3SO2R4; and R3, R4 are each        independently H, halogen, CH₂F, CHF₂, CF₃, C₁₋₄alkyl, C₁₋₄alkyl        cycloalkyl, C₁₋₄cyclohetalkyl, aryl, C₁₋₄ alkyl aryl, hetaryl,        C₁₋₄ alkyl hetaryl, or may be joined to form an optionally        substituted 3-8 membered (saturated or unsaturated) ring        optionally containing an atom selected from O, S, NR6; and R5 is        selected from H, C₁₋₄ alkyl, halogen, CH₂F, CHF₂, CF₃, aryl or        hetaryl; and R6 is selected from H, C₁₋₄ alkyl aryl, hetaryl,        C₁₋₄ alkyl aryl, C₁₋₄ alkyl hetaryl.

R1 is H, C₁₋₄alkyl, C₁₋₄ cycloalkyl.

Q is a bond, CH₂, C₁₋₄alkyl

A is aryl, hetaryl optionally substituted with 0-3 substituentsindependently chosen from halogen, C₁₋₄ alkyl, CH₂F, CHF₂, CF₃, OCF₃,CN, NR8R9, aryl, hetaryl, C₁₋₄aryl, C₁₋₄hetaryl, C₁₋₄ alkylNR8R9, OC₁₋₄alkylNR8R9, nitro, NR10C₁₋₄NR8R9, R8COR9, NR10CONR8R9, NR8SO₂R9,CONR8R9, CO₂R8 where R8 and R9 are each independently H, C₁₋₄ alkyl arylor which together form an optionally substituted 4-8 membered ring whichmay contain a heteroatom selected from O, S, NR11, where R11 is C₁₋₄alkyl, and R10 is selected from H, C₁₋₄alkyl.

W is selected from H, C₁₋₄alkyl, C₂₋₆alkenyl; where C₁₋₄alkyl orC₂₋₆alkenyl may be optionally substituted with C₁₋₄alkyl, OH,OC₁₋₄alkyl, NR12R13; and R12, and R13 are each independently H,C₁₋₄alkyl, or may be joined to form an optionally substituted 3-8membered ring optionally containing an atom selected from O, S, NR14 andR14 is selected from H, C₁₋₄ alkyl.

In the above description it will be appreciated that:

C₁₋₄ alkyl means a straight or branched alkyl chain

Aryl means unsubstituted or optionally substituted phenyl or naphthyl.

Hetaryl means a unsubstituted or optionally substituted 5- or 6-memberedheteroaromatic ring containing one or more heteroatoms selected from O,N, S.

Cycloalkyl means a 3-8 membered saturated ring

Cyclohetalkyl means a 3-8 membered saturated ring containing 1-3heteroatoms selected from O, S, NR13, where R13 is H, C₁₋₄ alkyl, aryl,hetaryl.

In a further preferred embodiment the compound is selected fromcompounds of the general formula II.

or pharmaceutically acceptable salts, hydrates, solvates, crystal formsor diastereomers thereof, wherein:

D is a heterocyclic ring selected from:

-   -   where X₁, X₂, X₃, X₄ are optionally substituted carbon, or one        of X₁, X₂, X₃, X₄ is N; R2 is 0-4 substituents independently        chosen from H, halogen, C₁₋₄ alkyl, CH₂F, CHF₂, CF₃, OCF₃, aryl,        hetaryl, C₁₋₄alkylOC₁₋₄alkyl, C₁₋₄alkylOaryl, C₁₋₄alkylNR3R-4,        CO₂R3, CONR3R4, CONR3SO₂R4, nitro, NR3R4, C₁₋₄ alkylNR3R4,        NR3COR4, NR5CONR3R4, NR3SO₂R4, C₁₋₄alkylNR3COR4,        C₁₋₄alkylNR5CONR3R4, C₁₋₄alkylNR3SO₂R4; and R3, R4 are each        independently H, halogen, CH₂F, CHF₂, CF₃, C₁₋₄ alkyl, C₁₋₄        alkyl cycloalkyl, C₁₋₄ cyclohetalkyl, aryl, C₁₋₄ alkyl aryl,        hetaryl, or may be joined to form an optionally substituted 3-8        membered (saturated or unsaturated) ring optionally containing        an atom selected from O, S, NR6; and R5 is selected from H, C₁₋₄        alkyl, halogen, CH₂F, CHF₂, CF₃, aryl or hetaryl; and R6 is        selected from H, C₁₋₄ alkyl, aryl, hetaryl, C₁₋₄, alkyl aryl,        C₁₋₄ alkyl hetaryl.

R1 is H, C₁₋₄ alkyl, C₁₋₆ cycloalkyl.

W is H, C₁₋₄ alkyl

A is aryl, hetaryl optionally substituted with 0-3 substituentsindependently chosen from halogen, C₁₋₄ alkyl, CH₂F, CHF₂, CF₃, OCF₃,CN, nitro, NR8R9, aryl, hetaryl, C₁₋₄aryl, C₁₋₄hetaryl, C₁₋₄ alkylNR8R9,OC₁₋₄ alkylNR8R9, NR10C₁₋₄NR8R9, NR8COR9, NR10CONR8R9, NR8SO₂R9,CONR8R9, CO₂R8 where R8 and R9 are each independently H, C₁₋₄ alkyl,aryl or which together form an optionally substituted 4-8 membered ringwhich may contain a heteroatom selected from O, S, NR11, where R11 isC₁₋₄ alkyl, and R10 is selected from H, C₁₋₄ alkyl.

In the above description it will be appreciated that:

C₁₋₄ alkyl means a straight or branched alkyl chain

Aryl means unsubstituted or optionally substituted phenyl or naphthyl.

Hetaryl means a unsubstituted or optionally substituted 5- or 6-memberedheteroaromatic ring containing one or more heteroatoms selected from O,N, S.

Cycloalkyl means a 3-8 membered saturated ring

Cyclohetalkyl means a 3-8 membered saturated ring containing 1-3heteroatoms selected from O, S, NR13, where R13 is H, C₁₋₄ alkyl, aryl,hetaryl.

The compounds of this invention include all conformational isomers (eg.cis and transisomers). The compounds of the present invention haveasymmetric centers and therefore exist in different enantiomeric anddiastereomeric forms. This invention relates to the use of all opticalisomers and stereoisomers of the compounds of the present invention, andmixtures thereof, and to all pharmaceutical compositions and methods oftreatment that may employ or contain them. In this regard, the inventionincludes both the E and Z configurations. The compounds of formula I mayalso exist as tautomers. This invention relates to the use of all suchtautomers and mixtures thereof.

This invention also encompasses pharmaceutical compositions containingprodrugs of compounds of the formula I. This invention also encompassesmethods of treating or preventing disorders that can be treated orprevented by the inhibition of protein kinases, such as JAK comprisingadministering prodrugs of compounds of the formula I. Compounds offormula I having free amino, amido, hydroxy or carboxylic groups can beconverted into prodrugs. Prodrugs include compounds wherein an aminoacid residue, or a polypeptide chain of two or more (eg, two, three orfour) amino acid residues which are covalently joined through peptidebonds to free amino, hydroxy and carboxylic acid groups of compounds offormula I. The amino acid residues include the 20 naturally occurringamino acids commonly designated by three letter symbols and alsoinclude, 4-hydroxyproline, hydroxylysine, demosine, isodemosine,3-methylhistidine, norvlin, beta-alanine, gamma-aminobutyric acid,citrulline, homocysteine, homoserine, ornithine and methioine sulfone.Prodrugs also include compounds wherein carbonates, carbamates, amidesand alkyl esters which are covalently bonded to the above substituentsof formula I through the carbonyl carbon prodrug sidechain. Prodrugsalso include phosphate derivatives of compounds of formula I (such asacids, salts of acids, or esters) joined through a phosphorus-oxygenbond to a free hydroxyl of compounds of formula I.

In a still further preferred embodiment the compound possesses Schirality at the chiral carbon bearing W, where W is C₁₋₄ alkyl. Thecompound can be used as a purified isomer or as a mixture of any ratioof isomers. It is however preferred that the mixture comprises at least70%, 80%, 90%, 95%, or 99% of the preferred isomer.

In a still further preferred embodiment the compound is selected fromthe group consisting of6-(1H-benzimidazol-1-yl)-N-[(1R)-1-phenylethyl]pyrazin-2-amine,N-benzyl-6-(1H-imidazol-1-yl)pyrazin-2-amine,6-(1H-benzimidazol-1-yl)-N-[(1S)-1-(4-methoxyphenyl)ethyl]pyrazin-2-amine,N-(4-Fluorobenzyl)-6-(1H-imidazol-1-yl)pyrazin-2-amine,6-(1H-benzimidazol-1-yl)-N-[(1S)-1-(4-bromophenyl)ethyl]pyrazin-2-amine,6-(1H-Imidazol-1-yl)-N-[(1R)-1-phenylethyl]pyrazin-2-amine,1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazole-6-carboxamide,6-(1H-Benzimidazol-1-yl)-N-benzylpyrazin-2-amine,1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazole-5-carboxamide,6-(1H-Benzimidazol-1-yl)-N-(4-fluorobenzyl)pyrazin-2-amine,6-{5-[(Morpholino-1-yl)carbonyl]-1H-benzimidazol-1-yl}-N-[(1S)-1-phenylethyl]pyrazin-2-amine,6-(1H-imidazo[4,5-b]pyridin-1-yl)-N-[(1R)-1-phenylethyl]pyrazin-2-amine,6-{6-[(Morpholino-1-yl)carbonyl]-1H-benzimidazol-1-yl}-N-[(1S)-1-phenylethyl]pyrazin-2-amine,6-(1H-imidazol-1-yl)-N-(4-morpholin-4-ylphenyl)pyrazin-2-amine,N-[1-(6-{[(1S)-1-Phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]cyclopropanecarboxamide,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]nicotinamide,N-[1-(6-{[(1S)-1-Phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]cyclopropanecarboxamide,6-(1H-Benzimidazol-1-yl)-N-[(1R)-1-phenylethyl]pyrazin-2-amine,6-[6-(4,5-dihydro-1,3-oxazol-2-yl)-1H-benzimidazol-1-yl]-N-[(1S)-1-phenylethyl]pyrazin-2-amine,N-[(1R)-1-Phenylethyl]-6-(4-phenyl-1H-imidazol-1-yl)pyrazin-2-amine,1-[6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl]-N-(2-hydroxyethyl)-1H-benzimidazole-6-carboxamide,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]methanesulfonamide,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]methanesulfonamide,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]isonicotinamide,6-(1H-Imidazol-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]isonicotinamide,6-(1H-Benzimidazol-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine,6-[5-(4,5-dihydro-1,3-oxazol-2-yl)-1H-benzimidazol-1-yl]-N-[(1S)-1-phenylethyl]pyrazin-2-amine,1-[6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl]-N-(2-hydroxyethyl)-1H-benzimidazole-5-carboxamide,6-(5-Methyl-1H-benzimidazol-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]nicotinamide,N-methyl-1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazole-5-carboxamide,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]2,2-dimethylpropanamide,N-methyl-1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazole-6-carboxamide,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]-2,2-dimethylpropanamide,1-(6-{[(1S)-1-Phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-amine,2-Methoxy-N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]acetamide,1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-amine,2-Methoxy-N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]acetamide,N-Benzyl-1-[6-([(1S)-1-phenylethyl]amino)pyrazin-2-yl]-1H-benzimidazole-5-carboxamide,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimindazol-5-yl]pyrazine-2-carboxamide,1-(6-{[(1S)-1-Phenylethyl]amino}pyrazin-2-yl)-N-phenyl-1H-benzimidazole-5-carboxamide,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]pyrazine-2-carboxamide,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]acetamide,6-{5-[(4-Methylpiperazin-1-yl)methyl]-1H-benzimidazol-1-yl}-N-[(1S)-1-phenylethyl]pyrazin-2-amine,N-[1-(6-{[(1S)-1-Phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]acetamide,[1-(6-{[(1S)-1-Phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]methanol,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]benzamide,[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]methanol,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]benzamide,1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-N-[2-(dimethylamino)ethyl]-1H-benzimidazole-5-carboxamide,1-[6-{[(1S)-1-Phenylethyl]amino}pyrazin-2-yl]-N-(pyridin-3-ylmethyl)-1H-benzimidazol-5-amine,tert-butyl(2S)-2-({[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]amino}carbonyl)pyrrolidine-1-carboxylate,6-(3H-imidazo[4,5-c]pyridin-3-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine,6-(1H-benzimidazol-1-yl)-N-[1-(4-fluorophenyl)ethyl]pyrazin-2-amine,6-(1H-imidazo[4,5-c]pyridin-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine,6-(1H-benzimidazol-1-yl)-N-[(1S)-1-(4-pyridin-3-ylphenyl)ethyl]pyrazin-2-amine,(2S)-N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]pyrrolidine-2-carboxamide,N-[(1S)-1-phenylethyl]-6-(5-pyridin-4-yl-1H-benzimidazol-1-yl)pyrazin-2-amine,N-[(1S)-1-phenylethyl]-6-(5-pyridin-3-yl-1H-benzimidazol-1-yl)pyrazin-2-amine,6-(5-bromo-1H-benzimidazol-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine,N-[3-(1H-imidazol-1-yl)propyl]-1-[6-([(1S)-1-phenylethyl]amino)pyrazin-2-yl]-1H-benzimidazole-6-carboxamide,N-1H-benzimidazole-6-carboxamide,N-(3-morpholin-4-ylpropyl)-1-[6-([(1S)-1-phenylethyl]amino)pyrazin-2-yl]-1H-benzimidazole-6-carboxamide,N-(3-morpholin-4-ylpropyl)-1-[6-([(1S)-1-phenylethyl]amino)pyrazin-2-yl]-1H-benzimidazole-5-carboxamide,N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]piperidine-3-carboxamide,6-(1H-benzimidazol-1-yl)-N-[(1S)-1-pyridin-3-ylethyl]pyrazin-2-amine,6-(1H-benzimidazol-1-yl)-N-[(1S)-1-(1,1′-biphenyl-4-yl)ethyl]pyrazin-2-amineN-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]benzenesulfonamideand6-(1H-benzimidazol-1-yl)-N-[(1S)-1-(1,1′-biphenyl-4-yl)ethyl]pyrazin-2-amine.

In a second aspect the present invention consists in a compositioncomprising a carrier and at least one compound of the first aspect ofthe invention.

In a third aspect the present invention consists in a method of treatinga protein kinase-associated disease state, the method comprisingadministering a therapeutically effective amount of at least onecompound of the first aspect of the invention or a therapeuticallyeffective amount of a composition of the second aspect of the invention.

In a preferred embodiment, the disease state involves a receptortyrosine kinase selected from the group consisting of EGF, HER2, HER3,HER4, IR, IGF-1R, IRR, PDGFR.alpha., PDGFR.beta., CSFIR, C-Kit,C-fms,Flk-1R, Flk4, KDR/Flk-1, Flt-1, FGFR-1R, FGFR-2R, FGFR-3R andFGFR-4R.

In another preferred embodiment, the disease state involves a cellulartyrosine kinase selected from the group consisting of Src, Frk, Btk,Csk, Abl, ZAP70, Fes/Fps, Fak, Ack, Yes, Fyn, Lyn, Lck, Blk, Hck, Fgrand Yrk.

In a further preferred embodiment, the disease state involves a tyrosinekinase selected from the group consisting of JAK1, JAK2, JAK3 and TYK2.

In a yet further preferred embodiment, the disease state involves aserine/threonine kinase selected from the group consisting of ERK2,c-Jun, p38 MAPK, PKA, PKB, PKC, a cyclin-dependent kinase, CDK1, CDK2,CDK3, CDK4, CDK5, CDK6, CDK7, CDK8, CDK9, CDK10, and CDK11.

In a preferred embodiment of the present invention the disease state isselected from the group consisting of Atopy, such as Allergic Asthma,Atopic Dermatitis (Eczema), and Allergic Rhinitis; Cell MediatedHypersensitivity, such as Allergic Contact Dermatitis andHypersensitivity Pneumonitis; Rheumatic Diseases, such as Systemic LupusErythematosus (SLE), Rheumatoid Arthritis, Juvenile Arthritis, Sjögren'sSyndrome, Scleroderma, Polymyositis, Ankylosing Spondylitis, PsoriaticArthritis; Other autoimmune diseases such as Type I diabetes, autoimmunethyroid disorders, and Alzheimer's disease; Viral Diseases, such asEpstein Barr Virus (EBV), Hepatitis B, Hepatitis C, HIV, HTLV 1,Varicella-Zoster Virus (VZV), Human Papilloma Virus (HPV), Cancer, suchas Leukemia, Lymphoma and Prostate Cancer.

As used herein the term “protein kinase-associated disease state” refersto those disorders which result from aberrant protein kinase activity,in particular JAK activity and/or which are alleviated by inhibition ofone or more of these enzymes. In further aspects the present inventionprovides the use of the compounds described in the preparation ofmedicaments for the treatment of protein kinase-associated diseasestates.

As used herein the term “JAK”, “JAK kinase” or “JAK family” refers toprotein tyrosine kinases which possess the characterizing features ofJAK1, JAK2, JAK3 and TYK as described herein.

The present invention provides pharmaceutical compositions comprising atleast one of the compounds of the present invention capable of treatinga protein kinase-associated disorder, such as a JAK associated disorder,in an amount effective therefor, and a pharmaceutically acceptablevehicle or diluent. The compositions of the present invention maycontain other therapeutic agents as described below, and may beformulated, for example, by employing conventional solid or liquidvehicles or diluents, as well as pharmaceutical additives of a typeappropriate to the mode of desired administration (for example,excipients, binders, preservatives, stabilizers, flavors, etc.)according to techniques such as those well known in the art ofpharmaceutical formulation.

The compounds of the present invention may be administered by anysuitable means, for example, orally, such as in the form of tablets,capsules, granules or powders; sublingually; buccally; parenterally,such as by subcutaneous, intravenous, intramuscular, or intracisternalinjection or infusion techniques (e.g., as sterile injectable aqueous ornon-aqueous solutions or suspensions); nasally such as by inhalationspray; topically, such as in the form of a cream or ointment; orrectally such as in the form of suppositories; in dosage unitformulations containing non-toxic, pharmaceutically acceptable vehiclesor diluents. The compounds may, for example, be administered in a formsuitable for immediate release or extended release. Immediate release orextended release may be achieved by the use of suitable pharmaceuticalcompositions comprising the present compounds, or, particularly in thecase of extended release, by the use of devices such as subcutaneousimplants or osmotic pumps. The compounds may also administeredliposomally.

In addition to primates, such as humans, a variety of other mammals canbe treated according to the method of the present invention. Forinstance, mammals including, but not limited to, cows, sheep, goats,horses, dogs, cats, guinea pigs, rats or other bovine, ovine, equine,canine, feline, rodent or murine species can be treated. However, themethod can also be practiced in other species, such as avian species(e.g., chickens).

Diseases and conditions associated with inflammation and infection canbe treated using the method of the present invention. In a preferredembodiment, the disease or condition is one in which the actions ofeosinophils and/or lymphocytes are to be inhibited or promoted, in orderto modulate the inflammatory response.

The subjects treated in the above methods, in whom which JAK inhibitionis desired, are mammals, including, but not limited to, cows, sheep,goats, horses, dogs, cats, guinea pigs, rats or other bovine, ovine,equine, canine, feline, rodent or murine species, and preferably a humanbeing, male or female.

The term “therapeutically effective amount” means the amount of thesubject composition that will elicit the biological or medical responseof a tissue, system, animal or human that is being sought by theresearcher, veterinarian, medical doctor or other clinician.

The term “composition” as used herein is intended to encompass a productcomprising the specified ingredients in the specified amounts, as wellas any product which results, directly or indirectly, from combinationof the specified ingredients in the specified amounts. By“pharmaceutically acceptable” it is meant the carrier, diluent orexcipient must be compatibles with the other ingredients of theformulation and not deleterious to the recipient thereof.

The terms “administration of” and or “administering a” compound shouldbe understood to mean providing a compound of the invention to theindividual in need of treatment.

The pharmaceutical compositions for the administration of the compoundsof this invention may conveniently be presented in dosage unit form andmay be prepared by any of the methods well known in the art of pharmacy.All methods include the step of bringing the active ingredient intoassociation with the carrier which constitutes one or more accessoryingredients. In general, the pharmaceutical compositions are prepared byuniformly and intimately bringing the active ingredient into associationwith a liquid carrier or a finely divided solid carrier or both, andthen, if necessary, shaping the product into the desired formulation. Inthe pharmaceutical composition the active object compound is included inan amount sufficient to produce the desired effect upon the process orcondition of diseases. As used herein, the term “composition” isintended to encompass a product comprising the specified ingredients inthe specified amounts, as well as any product which results, directly orindirectly, from combination of the specified ingredients in thespecified amounts.

The pharmaceutical compositions containing the active ingredient may bein a form suitable for oral use, for example, as tablets, troches,lozenges, aqueous or oily suspensions, dispersible powders or granules,emulsions, hard or soft capsules, or syrups or elixirs. Compositionsintended for oral use may be prepared according to any method known tothe art for the manufacture of pharmaceutical compositions and suchcompositions may contain one or more agents selected from the groupconsisting of sweetening agents, flavoring agents, coloring agents andpreserving agents in order to provide pharmaceutically elegant andpalatable preparations. Tablets contain the active ingredient inadmixture with non-toxic pharmaceutically acceptable excipients whichare suitable for the manufacture of tablets. These excipients may be forexample, inert diluents, such as calcium carbonate, sodium carbonate,lactose, calcium phosphate or sodium phosphate; granulating anddisintegrating agents, for example, corn starch, or alginic acid;binding agents, for example starch, gelatin or acacia, and lubricatingagents, for example magnesium stearate, stearic acid or talc. Thetablets may be uncoated or they may be coated by known techniques todelay disintegration and absorption in the gastrointestinal tract andthereby provide a sustained action over a longer period. For example, atime delay material such as glyceryl monostearate or glyceryl distearatemay be employed. They may also be coated to form osmotic therapeutictablets for control release.

Formulations for oral use may also be presented as hard gelatin capsuleswherein the active ingredient is mixed with an inert solid diluent, forexample, calcium carbonate, calcium phosphate or kaolin, or as softgelatin capsules wherein the active ingredient is mixed with water or anoil medium, for example peanut oil, liquid paraffin, or olive oil.

Aqueous suspensions contain the active materials in admixture withexcipients suitable for the manufacture of aqueous suspensions. Suchexcipients are suspending agents, for example sodiumcarboxymethylcellulose, methylcellulose, hydroxy-propylmethylcellulose,sodium alginate, polyvinyl-pyrrolidone, gum tragacanth and gum acacia;dispersing or wetting agents may be a naturally-occurring phosphatide,for example lecithin, or condensation products of an alkylene oxide withfatty acids, for example polyoxyethylene stearate, or condensationproducts of ethylene oxide with long chain aliphatic alcohols, forexample heptadecaethyleneoxycetanol, or condensation products ofethylene oxide with partial esters derived from fatty acids and ahexitol such as polyoxyethylene sorbitol monooleate, or condensationproducts of ethylene oxide with partial esters derived from fatty acidsand hexitol anhydrides, for example polyethylene sorbitan monooleate.The aqueous suspensions may also contain one or more preservatives, forexample ethyl, or n-propyl, p-hydroxybenzoate, one or more coloringagents, one or more flavoring agents, and one or more sweetening agents,such as sucrose or saccharin.

Oily suspensions may be formulated by suspending the active ingredientin a vegetable oil, for example arachis oil, olive oil, sesame oil orcoconut oil, or in a mineral oil such as liquid paraffin. The oilysuspensions may contain a thickening agent, for example beeswax, hardparaffin or cetyl alcohol. Sweetening agents such as those set forthabove, and flavoring agents may be added to provide a palatable oralpreparation. These compositions may be preserved by the addition of ananti-oxidant such as ascorbic acid.

Dispersible powders and granules suitable for preparation of an aqueoussuspension by the addition of water provide the active ingredient inadmixture with a dispersing or wetting agent, suspending agent and oneor more preservatives. Suitable dispersing or wetting agents andsuspending agents are exemplified by those already mentioned above.Additional excipients, for example sweetening, flavoring and coloringagents, may also be present.

The pharmaceutical compositions of the invention may also be in the formof oil-in-water emulsions. The oily phase may be a vegetable oil, forexample olive oil or arachis oil, or a mineral oil, for example liquidparaffin or mixtures of these. Suitable emulsifying agents may benaturally-occurring gums, for example gum acacia or gum tragacanth,naturally-occurring phosphatides, for example soy bean, lecithin andesters or partial esters derived from fatty acids and hexitolanhydrides, for example sorbitan monooleate, and condensation productsof the said partial esters with ethylene oxide, for examplepolyoxyethylene sorbitan monooleate. The emulsions may also containsweetening and flavoring agents.

Syrups and elixirs may be formulated with sweetening agents, for exampleglycerol, propylene glycol, sorbitol or sucrose. Such formulations mayalso contain a demulcent, a preservative and flavoring and coloringagents. The pharmaceutical compositions may be in the form of a sterileinjectable aqueous or oleagenous suspension. This suspension may beformulated according to the known art using those suitable dispersing orwetting agents and suspending agents which have been mentioned above.The sterile injectable preparation may also be a sterile injectablesolution or suspension in a non-toxic parenterally-acceptable diluent orsolvent, for example as a solution in 1,3-butane diol. Among theacceptable vehicles and solvents that may be employed are water,Ringer's solution and isotonic sodium chloride solution. In additionssterile, fixed oils are conventionally employed as a solvent orsuspending medium. For this propose any bland fixed oil may be employedincluding synthetic mono- or diglycerides. In addition, fatty acids suchas oleic acid find use in the preparation of injectables.

The compounds of the present invention may also be administered in theform of suppositories for rectal administration of the drug. Thesecompositions can be prepared by mixing the drug with a suitablenon-irritating excipient which is solid at ordinary temperatures butliquid at the rectal temperature and will therefore melt in the rectumto release the drug. Such materials are cocoa butter and polyethyleneglycols.

For topical use, creams, ointments, jellies, solutions or suspensions,etc., containing the compounds of the present invention are employed.(For purposes of this application, topical application shall includemouthwashes and gargles.)

The compounds of the present invention can also be administered in theform of liposomes. As is known in the art, liposomes are generallyderived from phospholipids or other lipid substances. Liposomes areformed by mono- or multilamellar hydrated liquid crystals that aredispersed in an aqueous medium. Any non-toxic, physiologicallyacceptable and metabolisable lipid capable of forming liposomes can beused. The present compositions in liposome form can contain, in additionto a compound of the present invention, stabilisers, preservatives,excipients and the like. The preferred lipids are the phospholipids andphosphatidyl cholines, both natural and synthetic. Methods to formliposomes are known in the art.

The pharmaceutical composition and method of the present invention mayfurther comprise other therapeutically active compounds as noted hereinwhich are usually applied in the treatment of the above mentionedpathological conditions. Selection of the appropriate agents for use incombination therapy may be made by one of ordinary skill in the art,according to conventional pharmaceutical principles. The combination oftherapeutic agents may act synergistically to effect the treatment orprevention of the various disorders described above. Using thisapproach, one may be able to achieve therapeutic efficacy with lowerdosages of each agent, thus reducing the potential for adverse sideeffects.

Examples of other therapeutic agents include the following: cyclosporins(e.g., cyclosporin A), CTLA4-Ig, antibodies such as ICAM-3, anti-IL-2receptor (Anti-Tac), anti-CD45RB, anti-CD2, anti-CD3 (OKT-3), anti-CD4,anti-CD80, anti-CD86, agents blocking the interaction between CD40 andgp39, such as antibodies specific for CD40 and/or gp39 (i.e., CD154),fusion proteins constructed from CD40 and gp39 (CD401 g and CD8gp39),inhibitors, such as nuclear translocation inhibitors, of NF-kappa Bfunction, such as deoxyspergualin (DSG), cholesterol biosynthesisinhibitors such as HMC CoA reductase inhibitors (lovastatin andsimvastatin), non-steroidal antiinflammatory drugs (NSAIDs) such asibuprofen, aspirin, acetaminophen and cyclooxygenase inhibitors such asrofecoxib, steroids such as prednisolone or dexamethasone, goldcompounds, antiproliferative agents such as methotrexate, FK506(tacrolimus, Prograf), mycophenolate mofetil, cytotoxic drugs such asazathioprine, VP-16, etoposide, fludarabine, cisplatin andcyclophosphamide, TNF-α inhibitors such as tenidap, anti-TNF antibodiesor soluble TNF receptor, and rapamycin (sirolimus or Rapamune) orderivatives thereof.

When other therapeutic agents are employed in combination with thecompounds of the present invention they may be used for example inamounts as noted in the Physician Desk Reference (PDR) or as otherwisedetermined by one of ordinary skill in the art.

In the treatment or prevention of conditions which require proteintyrosine kinase inhibition an appropriate dosage level will generally beabout 0.01 to 500 mg per kg patient body weight per day which can beadministered in single or multiple doses. Preferably, the dosage levelwill be about 0.1 to about 250 mg/kg per day; more preferably about 0.5to about 100 mg/kg per day. A suitable dosage level may be about 0.01 to250 mg/kg per day, about 0.05 to 100 mg/kg per day, or about 0.1 to 50mg/kg per day. Within this range the dosage may be 0.05 to 0.5, 0.5 to 5or 5 to 50 mg/kg per day. For oral administration, the compositions arepreferably provided in the form of tablets containing 1.0 to 1000milligrams of the active ingredient, particularly 1.0, 5.0, 10.0, 15.0.20.0, 25.0, 50.0, 75.0, 100.0, 150.0, 200.0, 250.0, 300.0, 400.0, 500.0,600.0, 750.0, 800.0, 900.0, and 1000.0 milligrams of the activeingredient for the symptomatic adjustment of the dosage to the patientto be treated. The compounds may be administered on a regimen of 1 to 4times per day, preferably once or twice per day.

It will be understood, however, that the specific dose level andfrequency of dosage for any particular patient may be varied and willdepend upon a variety of factors including the activity of the specificcompound employed, the metabolic stability and length of action of thatcompound, the age, body weight, general health, sex; diet mode and timeof administration, rate of excretion, drug combination, the severity ofthe particular condition, and the host undergoing therapy.

Throughout this specification the word “comprise”, or variations such as“comprises” or “comprising”, will be understood to imply the inclusionof a stated element, integer or step, or group of elements, integers orsteps, but not the exclusion of any other element, integer or step, orgroup of elements, integers or steps.

All publications mentioned in this specification are herein incorporatedby reference.

Any discussion of documents, acts, materials, devices, articles or thelike which has been included in the present specification is solely forthe purpose of providing a context for the present invention. It is notto be taken as an admission that any or all of these matters form partof the prior art base or were common general knowledge in the fieldrelevant to the present invention as it existed in Australia before thepriority date of each claim of this application.

In order that the nature of the present invention may be more clearlyunderstood preferred forms thereof will now be described by reference tothe following non-limiting Examples.

Materials and Methods:

Compound Synthesis

Compounds are generally prepared in a 2-step process starting from2,6-dichloropyrazine.

The first step is a nucleophilic aromatic substitution to generate amonoamino-monohalo intermediate. (Scheme 1).

The nucleophilic aromatic substitution is typically carried out byaddition of a primary amine to the di-halogenated heterocycle in asolvent such as ethanol, isopropanol, tert-butanol, dioxane, THF, DMF,toluene or xylene. The reaction is typically performed at elevatedtemperature in the presence of excess amine or a non-nucleophilic basesuch as triethylamine or diisopropylethylamine, or an inorganic basesuch as potassium carbonate or sodium carbonate.

Alternatively, the amino substituent may be introduced through atransition metal catalysed amination reaction. Typical catalysts forsuch transformations include Pd(OAc)₂/P(t-Bu)₃, Pd₂(dba)₃/BINAP andPd(OAc)₂/BINAP.

The amines employed in the first step of the synthesis of thesecompounds are obtained commercially or are prepared using methods wellknown to those skilled in the art. Of particular interest areα-methylbenzylamines which are obtained commercially or may be preparedthrough reduction of oximes (Scheme 2). Typical reductants includelithium aluminium hydride, hydrogen gas in the presence of catalyticpalladium on charcoal, Zn in the presence of hydrochloric acid, sodiumborohydride in the presence of a Lewis acid such as TiCl₃, ZrCl₄, NiCl₂and MoO₃, or sodium borohydride in conjunction with Amberlyst H15 ionexchange resin and LiCl. The oximes are obtained in one-step from thecorresponding ketones through condensation with hydroxylamine. Thisreaction is generally performed in a protic solvent such as water orethanol, at temperatures from 0° C. to reflux. The hydroxylamine isgenerally used in the form of its hydrochloride salt, and therefore thereaction is performed in the presence of a base such as sodiumhydroxide. The ketones employed as starting materials are generallyobtained commercially or via procedures well known to those skilled inthe art.

α-Methyl benzylamines of high optical purity may be prepared from chiralα-methyl benzyl alcohols using methods well known to those skilled inthe art. Such methods include derivatisation of the hydroxyl as amesylate or tosylate and displacement with a nitrogen nucleophile, suchas phthalimide or azide which can then converted to the primary amineusing conventional synthetic methods; or, displacement of the hydroxylwith a suitable nitrogen nucleophile under Mitsunobu conditions. Thechiral α-methyl benzyl alcohols may be obtained through chiral reductionof the corresponding ketones. Chiral reducing methods are now well knownin organic chemistry and include enzymatic processes, asymmetrichydrogenation procedures and chiral oxazaborolidines.

The second step of the synthesis involves a nucleophilic aromaticsubstitution reaction of the monochloro- mono-amino pyrazine withimidazole, benzimidazole or indazole. The reaction is typicallyperformed using a salt of the imidazole, benzimidazole or indazole insolvents such as tetrahydrofuran, dimethylformamide, toluene, or xylenefrom room temperature toe reflux. The imidazole, benzimidazole orindazole salt is prepared by reaction with a metal hydride such assodium or potassium hydride or by reaction with caesium carbonate.Alternatively, a metal-catalysed coupling reaction can be used tointroduce the imidazole, benzimidazole or indazole ring. The reaction istypically performed using a base such as caesium carbonate, rubidiumcarbonate, potassium carbonate, sodium tert-butoxide or potassiumphosphate in a solvent such as xylene, toluene, and DMF from roomtemperature to reflux. Auxiliary reagents such as phase transfer agents(e.g. cetrimonium bromide) or copper complexing agents (e.g.phenanthroline) may also be employed in the reaction.

The imidazole, benzimidazole or indazole components used in thisreaction are obtained commercially or are prepared from commerciallyavailable imidazoles, benzimidazoles or indazoles via techniques wellknown to those skilled in the art.

Alternatively, an imidazole, benzimidazole or indazole derivative may bereacted with the mono-amino mono-chloro pyrazine and the subsequentproduct further derivatised using methods well known to those skilled inthe art.

Representative syntheses are reported below.

EXAMPLE 1 6-Chloro-N-[(1R)-1-phenylethyl]pyrazin-2-amine

A solution of R-α-methylbenzylamine (3.64 g, 30.0 mmol) and2,6-dichloropyrazine (1.50 g, 10.0 mmol) in dioxane (5 mL) was heated atreflux under N₂ for 48 hours. The solvent was removed and the productcrystallised from toluene-hexane.

¹H-n.m.r. (CDCl₃) δ1.59 (d, 3H, J=6.9 Hz, CH₃), 4.88 (q, 1H, J=6.6 Hz,CH), 5.13 (br s, 1H, NH), 7.27-7.36 (m, 5H, ArH), 7.64 (s, 1H, pyraz-H),7.79 (s, 1H, pyraz-H).

EXAMPLE 2 6-(1H-Imidazol-1-yl)-N-[(1R)-1-phenylethyl]pyrazin-2-amine

To a stirred solution of imidazole (82 mg 1.2 mmol) in anhydrous DMF (5mL) at 0° C. under N₂ was added sodium hydride (58 mg, 60 dispersion inoil, 1.45 mmol). The mixture was stirred at 0° C. for 10 min and at RTfor 30 min. To this mixture was added a solution of6-chloro-N-[(1R)-1-phenylethyl]pyrazin-2-amine (280 mg, 1.2 mmol) in DMF(5 mL). The mixture was then stirred at RT for 62 h followed by heatingat reflux for 18 h. The DMF was removed under reduced pressure and theresidue diluted with chloroform. The organic layer was washed withwater, dried (Na₂SO₄) and the solvent removed under reduced pressure tofurnish the crude product. Column chromatography usingdichloromethane-methanol (19:1→10.1) as eluant separated recoveredstarting material from product (177 mg, 56%).

¹H-n.m.r. (CDCl₃) δ1.62 (d, 3H, J=6.9 Hz, CH₃), 4.97 (m, 1H, CH), 5.46(d, 1H, J=6.0 Hz, NH), 7.17 (s, 1H, imid-H), 7.23-7.40 (m, 5H, Ph-H),7.47 (s, 1H, imid-H), 7.76 (s, 1H, pyraz-H), 7.91 (s, 1H, pyraz-H), 8.20(s, 1H, imid-H).

m/z (ES) 266 (M⁺+H), 162, 105.

EXAMPLE 3 6-Chloro-N-(4-morpholin-4-ylphenyl)pyrazin-2-amine

In a procedure analogous to example 1, reaction of 4-morpholinoaniline(2.15 g, 12.1 mmol) and 2,6-dichloropyrazine (0.756 g, 5.03 mmol)furnished the product (0.54 g, 37%).

¹H-n.m.r. (CDCl₃) δ3.25 (br s, 4H, CH₂), 3.99 (br s, 4H, CH₂), 7.05-7.17(m, 2H, ArH), 7.42-7.54 (m, 2H, ArH), 7.94 (s, 1H, pyraz-H), 8.04 (s,1H, pyraz-H), 8.06 (s, 1H, NH).

EXAMPLE 4 6-Chloro-N-(4-fluorobenzyl)pyrazin-2-amine

In a procedure analogous to example 1, reaction of 4-fluorobenzylamine(3.75 g, 30.0 mmol) and 2,6-dichloropyrazine (1.49 g, 10.0 mmol)furnished the product (2.35 g, 99%).

¹H-n.m.r. (CDCl₃) δ 4.53(s, 2H, CH₂), 5.08(br s, 1H, NH), 7.01-7.07(m,2H, ArH), 7.30-7.34(m, 2H, ArH), 7.77(s, 1H, pyraz-H), 7.83(s, 1H,pyraz-H)

EXAMPLE 5 6-(1H-Imidazol-1-yl)-N-(4-morpholin-4-ylphenyl)pyrazin-2-amine

In a procedure analogous to example 2, reaction of6-chloro-N-(4-morpholin-4-ylphenyl)pyrazin-2-amine (100 mg, 0.34 mmol)and imidazole (126 mg, 0.38 mmol) furnished the product (37 mg, 34%).

¹H-n.m.r. (CDCl₃) δ3.18 (br s, 4H, 2CH₂), 3.87-3.91 (m, 4H, 2CH₂), 6.77(s, 1H, imid-H), 6.94-6.98 (m, 2H, ArH), 7.36 (d, 2H, J=8.7 Hz, ArH),7.62 (br s, 1H, imid-H), 8.03 (s, 1H, pyraz-H), 8.08 (s, 1H, pyraz-H),8.39 (br s, 1H, imid-H).

m/z (ES) 323 (M⁺+H).

EXAMPLE 6 N-(4-Fluorobenzyl)-6-(H-imidazol-1-yl)pyrazin-2-amine

In a procedure analogous to example 2, reaction of6-chloro-N-(4-fluorobenzyl)pyrazin-2-amine (240 mg, 1.01 mmol) andimidazole (76 mg, 1.11 mmol) furnished the product (210 mg, 65%).

¹H-n.m.r. (CDCl₃) δ 4.59 (d, 2H, J=5.7 Hz, CH₂), 5.23 (t-like, 1H, NH),7.03-7.08 (m, 2H, ArH), 7.20 (s, 1H, imid-H), 7.32-7.37 (m, 2H, ArH),7.55 (s, 1H, imid-H), 7.85 (s, 1H, pyraz-H), 8.00 (s, 1H, pyraz-H), 8.29(s, 1H, imid-H).

m/z (ES)270 (M⁺+H).

EXAMPLE 76-(2-Methyl-1H-imidazol-1-yl)-N-[(1R)-1-phenylethyl]pyrazin-2-amine

In a procedure analogous to example 2, reaction of6-chloro-N-[(1R)-1-phenylethyl]pyrazin-2-amine (150 mg, 0.64 mmol) and2-methylimidazole (58 mg, 0.71 mmol) furnished the product (172 mg,40%).

¹H-n.m.r. (CDCl₃) δ1.59 (d, 3H, J=6.8 Hz, CH₃), 2.43 (s, 3H, CH₃), 4.98(m, 1H, CH), 5.45 (br s, 1H, NH), 6.98 (d, 1H, J=1.3 Hz, imid-H), 7.17(d, 1H, J=1.3 Hz, imid-H), 7.22-7.35 (m, 5H, ArH), 7.82 (s, 1H,pyraz-H), 7.84 (s, 1H, pyraz-H).

m/z (ES) 280 (M⁺+H).

EXAMPLE 8N-(4-Fluorobenzyl)-6-(2-methyl-1H-imidazol-1-yl)pyrazin-2-amine

In a procedure-analogous to example 2 reaction of6-chloro-N-(4-fluorobenzyl)pyrazin-2-amine (150 mg, 0.63 mmol) and2-methylimidazole (57 mg, 0.69 mmol) furnished the product (42 mg, 23%).

¹H-n.m.r. (CDCl₃) δ2.56 (s, 3H, CH₃), 4.57 (d, J=5.7 Hz, 1H, CH₂), 5.34(br s, 1H, NH), 7.01-7.07 (m, 3H, Ar—H), 7.26 (s, 1H, imid-H), 7.29-7.34(m, 2H, ArH), 7.92 (s, 2H, pyraz-H).

m/z (ES) 284 (M⁺+H).

EXAMPLE 9N-(4-Fluorobenzyl)-6-(4-methyl-1H-imidazol-1-yl)pyrazin-2-amine andN-(4-fluorobenzyl)-6-(5-methyl-1H-imidazol-1-yl)pyrazin-2-amine

In a procedure analogous to example 2, reaction of2-(4-fluorobenzylamino)-6-chloro-pyrazine (190 mg, 0.80 mmol) and4-methylimidazole (72 mg, 0.88 mmol) furnished the following products:4-methyl derivative (100 mg, 44%); 5-methyl derivative (19 mg, 8%).

(4-methyl derivative) ¹H-n.m.r. (CDCl₃) δ 2.31 (s, 3H, CH₃), 4.58 (d,2H, J=5.7 Hz, CH₂), 5.40 (br s, 1H, NH), 6.92 (s, 1H, imid-H), 7.00-7.08(m, 2H, ArH), 7.25 (s, 1H, imid-H5), 7.31-7.36 (m, 2H, ArH), 7.84 (s,1H, pyraz-H5), 7.93 (s, 1H, pyraz-H3), 8.24 (br s, 1H, imid-H2).

m/z (ES) 284 (M⁺+H).

(5-methyl derivative) ¹H-n.m.r. (CDCl₃) δ 2.34 (s, 3H, CH₃), 4.57 (d,2H, J=5.7 Hz, CH₂), 5.44 (br s, 1H, NH), 6.92 (s, 1H, imid-H), 7.00-7.07(m, 2H, ArH), 7.28-7.34 (m, 2H, ArH), 7.93 (s, 1H, pyraz-H), 7.95 (s,1H, pyraz-H), 7.98 (br s, 1H, imid-H).

m/z (ES) 284 (M⁺+H).

EXAMPLE 10N-[(1R)-1-Phenylethyl]-6-(4-phenyl-1H-imidazol-1-yl)pyrazin-2-amine

In a procedure analogous to example 2, reaction of6-chloro-N-[(1R)-1-phenylethyl]pyrazin-2-amine (180 mg, 0.77 mmol) and4-phenylimidazole (122 mg, 0.85 mmol) furnished the product (176 mg,67%).

¹H-n.m.r. (CDCl₃) δ 1.63 (d, 3H, J=6.9 Hz, CH₃), 4.93-5.02 (m, 1H, CH),5.26 (d, 1H, J=6.0 Hz, NH), 7.25-7.44 (m, 8H, ArH), 7.72 (d, 1H, J=1.2Hz, imid-H), 7.77 (s, 1H, pyraz-H), 7.82-7.86 (m, 2H, ArH), 7.92 (s, 1H,pyraz-H), 8.22 (s, 1H, imid-H).

m/z (ES) 342 (M⁺+H).

EXAMPLE 11 N-Benzyl-6-chloropyrazin-2-amine

In a procedure analogous to example 1, reaction of benzylamine (3.21 g,30.0 mmol) and 2,6-dichloropyrazine (1.49 g, 10.0 mmol) furnished theproduct (2.15 g, 98%).

¹H-n.m.r. (CDCl₃) δ 4.55 (d, 2H, J=5.7 Hz, CH₂), 7.28-7.40 (m, 5H, ArH),7.76 (s, 1H, pyraz-H), 7.83 (s, 1H, pyraz-H).

EXAMPLE 12 6-(1H-Benzimidazol-1-yl)-N-benzylpyrazin-2-amine

To a stirred solution of benzimidazole (130 mg, 1.1 mmol) in anhydrousDMF (5 mL) at 0° C. under N₂ was added sodium hydride (56 mg, 60%dispersion in oil, 1.45 mmol) in portions over 2 min. The mixture wasstirred at 0° C. for 15 min and at RT for 60 min. To this was added asolution of (6-chloro-pyrazin-2-yl)-(1-benzyl)-amine (220 mg) in DMF (5mL) and the resulting mixture was then heated at reflux for 18 h. TheDMF was removed under reduced pressure and the residue diluted withchoroform. The organic layer was washed with water, dried (Na₂SO₄) andthe solvent removed under reduced pressure to furnish the crude product.Column chromatography using dichloromethane-methanol (20:1→10:1) aseluant separated the product (100 mg).

¹H-n.m.r. (CDCl₃) δ 4.66 (d, 2H, J=5.7 Hz, CH₂), 5.56 (m, 1H, NH),7.29-7.39 (m, 7H, Ar—H), 7.78-7.89 (m, 2H, Ar—H), 7.92 (s, 1H, pyraz-H),8.16 (s, 1H, pyraz-H), 8.48 (s, 1H, benzimid-H2).

m/z (ES) 302 (M⁺+H).

EXAMPLE 136-(1H-Benzimidazol-1-yl)-N-[(1R)-1-phenylethyl]pyrazin-2-amine

In a procedure analogous to example 12, reaction of6-chloro-N-[(1R)-1-phenylethyl]pyrazin-2-amine (240 mg, 1.03 mmol) andbenzimidazole (130 mg; 1.10 mmol) furnished the product (187 mg, 59%).

¹H-n.m.r. (CDCl₃) δ1.63 (d, 3H, J=6.6 Hz, CH₃), 4.98-5.20 (m, 1H, CH),5.58 (d, 1H, J=6.0 Hz, NH), 7.25-7.42 (m, 6H, Ph-H, benzimid-H), 7.70(dd, 1H, J=7.2, 1.0 Hz, benzimid-H), 7.82 (dd, 1H, J=8.0, 1.2 Hz,benzimid-H), 7.87 (s, 1H, pyraz-H), 8.11 (s, 1H, pyraz-H), 8.38 (s, 1H,benzimid-H).

m/z (ES) 315 (M⁺+H), 212, 105.

EXAMPLE 146-(1H-Benzimidazol-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine

In a procedure analogous to example 12, reaction of6-chloro-N-[(1S)-1-phenylethyl]pyrazin-2-amine (140 mg, 0.60 mmol) andbenzimidazole (78 mg, 0.66 mmol) furnished the product (71 mg, 38%).

¹H-n.m.r. (CDCl₃) δ1.57 (d, 3H, J=6.9 Hz, CH₃), 4.95 (m, 1H, CH), 5.29(d, 1H, J=6.0 Hz, NH), 7.19-7.35 (m, 7H, Ph-H, benzimid-H), 7.63-7.66(m, 1H, benzimid-H), 7.74-7.77 (m, 1H, benzimid-H), 7.78 (s, 1H,pyraz-H), 8.06 (s, 1H, pyraz-H), 8.31 (s, 1H, benzimid-H).

m/z (ES) 316 (M⁺+H), 212, 105

EXAMPLE 156-(1H-Benzimidazol-1-yl)-N-(4-morpholin-4-ylphenyl)pyrazin-2-amine

In a procedure analogous to example 12, reaction of6-chloro-N-(4-morpholin-4-ylphenyl)pyrazin-2-amine (150 mg, 0.52 mmol)and benzimidazole (67 mg, 0.57 mmol) furnished the product (60 mg, 31%).

¹H-n.m.r. (CDCl₃) δ 3.19 (br s, 4H, 2CH₂), 3.90 (t, 4H, J=4.6 Hz, 2CH₂),6.69 (s, 1H, NH), 6.98 (d, 2H, J=8.4 Hz, ArH), 7.37-7.41 (m, 4H, ArH),7.87-7.90 (m, 1H, ArH), 8.00-8.03 (m, 1H, ArH), 8.08 (s, 1H, pyraz-H),8.31 (s, 1H, pyraz-H), 8.59 (s, 1H, benzimid-H2).

m/z-(ES) 373 (M⁺+H).

EXAMPLE 16 6-(1H-Benzimidazol-1-yl)-N-(4-fluorobenzyl)pyrazin-2-amine

In a procedure analogous to example 12, reaction of6-chloro-N-(4-fluorobenzyl)pyrazin-2-amine (240 mg, 1.01 mmol) andbenzimidazole (130 mg, 1.1 mmol) furnished the product (170 mg, 53%).

¹H-n.m.r. (CDCl₃) δ4.64 (d, 2H, J=5.7 Hz, CH₂), 5.46 (br s, 1H, NH),7.06 (m, 2H, ArH), 7.30-7.38 (m, 4H, ArH), 7.82-7.88 (m, 2H, ArH), 7.93(s, 1H, pyraz-H), 8.20 (s, 1H, benzimid-H), 8.49 (s, 1H, pyraz-H).

m/z (ES) 320 (M⁺+H).

EXAMPLE 176-(1H-imidazo[4,5-b]pyridin-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amineand 6-(3H-imidazo[4,5-b]pyridin3-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine

In a procedure analogous to example 12, reaction of6-chloro-N-[(1S)-1-phenylethyl]pyrazin-2-amine (240 mg, 1.03 mmol) and4-azabenzimidazole (130 mg, 1.09 mmol) furnished the product (7 mg, 2%)as a 1:1 mixture of regioisomers.

¹H-n.m.r. (as 1:1 mixture) (CDCl₃) δ1.54 (d, 3H, CH₃), 1.63 (d, 3H,CH₃), 4.63 (br s, 1H, NH), 4.82-4.91 (m, 1H, CH), 4.95-5.04 (m, 1H, NH),5.16 (m, 1H, NH), 7.07 (s, 1H, ArH), 7.22-7.43 (m, 16H, ArH), 7.87 (s,1H, ArH), 8.11 (dd, 1H, J=8.1, 1.5 Hz, ArH), 8.48 (dd, 1H, J=4.8, 1.2Hz, ArH), 8.80 (s, 1H, ArH).

m/z (ES) 317 (M⁺+H).

EXAMPLE 186-(5-Methyl-1H-benzimidazol-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amineand6-(6-methyl-1H-benzimidazol-1-yl)-N-[(1S)-phenylethyl]pyrazin-2-amine

In a procedure analogous, to example 12, reaction of6-chloro-N-[(1S)-1-phenylethyl]pyrazin-2-amine (91 mg, 0.39 mmol) and5-methylbenzimidazole (57 mg, 0.43 mmol) furnished the product (61 mg,148%) as a 1:1 mixture.

¹H-n.m.r. (as 1:1 mixture) (CDCl₃) δ1.65 (d, 3H, CH₃), 1.66 (d, 3H,CH₃), 2.49 (s, 3H, CH₃), 2.50 (s, 3H, CH₃), 5.01 (m, 1H, CH), 5.04 (m,1H, CH), 5.56 (d, 1H, NH), 5.62 (d, 1H, NH), 7.27-7.45 (m, 10H, Ph-H),7.56-7.85 (m, 4H, benzimid-H), 7.94 (s, 1H, pyraz-H), 7.97 (s, 1H,pyraz-H), 8.02 (s, 2H, benzimid-H), 8.16 (s, 1H, pyraz-H), 8.17 (s, 1H,pyraz-H), 8.59 (s, 1H, benzimid-H), 8.70 (s, 1H, benzimid-H).

m/z (ES) 330 (M⁺+H).

EXAMPLE 196-(2-Methyl-1H-benzimidazol-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine

In a procedure analogous to example 12, reaction of6-chloro-N-[(1S)-1-phenylethyl]pyrazin-2-amine (120 mg, 0.51 mmol) and2-methyl-benzimidazole (75 mg, 0.57 mmol) furnished the product (18 mg,11%).

¹H-n.m.r. (CDCl₃) δ1.63 (d, 3H, J=6.9 Hz, CH₃), 2.59 (s, 3H, CH₃), 4.93(m, 1H, CH), 5.89 (br s, 1H, NH), 7.16-7.49 (m, 7H, ArH), 7.93 (d, 2H,J=8.1 Hz, Ar—H), 8.00 (br s, 1H, pyraz-H), 8.23 (br s, 1H, pyraz-H).

m/z (ES) 330 (M⁺+H).

EXAMPLE 20 N-(2-Hydroxyethyl)-1H-benzimidazole-5-carboxamide

To a stirred suspension of benzimidazole-5-carboxylic acid (0.40 g, 2.50mmol) in benzene (8 mL) was added thionyl chloride (2 mL) dropwise atroom temperature. The whole mixture was then heated under reflux for 5h. Benzene and thionyl chloride was evaporated off under reducedpressure, and the obtained acid chloride was suspended intetrohydrofuran. To this was the added 2-hydroxyethyl amine dropwise at0° C., and the resultant mixture was then stirred at room temperatureovernight. The solvent was then decanted, and the residue was washedwith diethyl ether (40 mL). The residue oil was then treated with coldwater (5 mL), and the aqueous solution was carefully decanted. This stepwas repeated, and now the amide was obtained as a pale brown solid, 0.19g (37%).

¹H-n.m.r. (CDCl₃) δ 3.30-3.37 (m, 2H, CH₂), 3.51-3.55 (m, 2H, CH₂), 4.69(s, 1H, OH), 7.59 (d, 1H, J=8.1 Hz, ArH), 7.73 (d, 1H, J=8.4 Hz, ArH),8.13 (br s, 1H, NH), 8.30 (s, 1H, H-4), 8.35 (br s, 1H, H-2), 12.47 (brs, 1H, CONH).

EXAMPLE 211-[6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl]-N-(2-hydroxyethyl)-1H-benzimidazole-5-carboxamide

In a procedure analogous to example 12, reaction of6-chloro-N-[(1S)-1-phenylethyl]pyrazin-2-amine (91 mg, 0.39 mmol) andbenzimidazole 5-carboxylic acid (2-hydroxyethyl)amide (88 mg, 0.43 mmol)furnished the product which was purified by column chromatography (22mg, 14%).

¹H-n.m.r. (CDCl₃) δ1.63 (d, 3H, J=6.9 Hz, CH₃), 3.68 (2H, dt, J=5.1 Hz,CH₂NH), 3.91 (2H, t, J=5.1 Hz, CH₂OH), 4.98 (m, 1H, CH), 5.50 (d, 1H,J=6.0 Hz, NH), 7.15 (1H, t, J=5.1 Hz, CONH), 7.28-7.41 (m, 5H, Ar—H),7.60 (d, 1H, J=8.4 Hz, benzimid-H), 7.73 (d, 1H, J=8.4 Hz, benzimid-H),7.88 (s, 1H, pyraz-H), 8.05 (s, 1H, pyraz-H), 8.20 (s, 1H, benzimid-H),8.40 (s, 1H, benzimid-H). m/z (ES) 403 (M⁺+H).

EXAMPLE 22[1-(6-{[1S)-1-Phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]methanoland[1-(6{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]methanol

In a procedure analogous to example 12, reaction of6-chloro-N-[(1S)-1-phenylethyl]pyrazin-2-amine (1.07 g, 4.6 mmol) and5-hyroxymethyl benzimidazole (0.68 g, Y 4.60 mmol) furnished the twoproducts separated by column chromatography: 5-hyroxymethyl isomer (34mg, 2%); 6-hydroxymethyl isomer (36 mg, 2%).

(5-hydroxymethyl isomer) ¹H-n.m.r. (CDCl₃) δ1.65 (d, 3H, J=6.9 Hz, CH₃),4.82 (s, 2H, CH₂OH), 5.03 (m, 1H, CH), 5.26 (d, 1H, J=6.0 Hz, NH),7.25-7.40 (m, 6H, Ar—H), 7.68 (d, 1H, J=8.4 Hz, benzimid-H), 7.81 (s,1H, benzimid-H), 7.85 (s, 1H, pyraz-H), 8.12 (s, 1H, pyraz-H), 8.39 (s,1H, benzimid-H). m/z (ES) 346 (M⁺+H).

(6-hydroxymethyl isomer) ¹H-n.m.r. (CDCl₃) δ1.63 (d, 3H, J=609 Hz, CH₃),4.79 (s, 2H, CH₂OH), 5.03 (m, 1H, CH), 5.42 (d, 1H, J=6.3 Hz, NH),7.25-7.42 (m, 6H, Ar—H), 7.78 (d, 1H, J=8.4 Hz, benzimid-H), 7.81 (s,1H, pyraz-H), 7.90 (s, 1H, benzimid-H), 8.10 (s, 1H, pyraz-H), 8.35 (s,1H, benzimid-H). m/z (ES) 346 (M⁺+H).

EXAMPLE 23 N-Methyl-1H-benzimidazole-5-carboxamide

In a procedure analogous to example 20, reaction ofbenzimidazole-5-carboxylic acid and aqueous methylamine furnished theproduct in 67% yield.

¹H-n.m.r. (CDCl₃) δ 2.79 (s, 3H, CH₃), 7.59 (d, 1H, J=8.4 Hz, H-6), 7.71(dd, 1H, J=8.4, 1.2 Hz, H-7), 8.10 (s, 1H, H-4), 8.29 (s, 1H, H-2), 8.36(br s, 1H, NH), 12.56 (br s, 1H, CONH).

EXAMPLE 24N-Methyl-1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazole-5-carboxamideand1-N-Methyl-1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazole-6-carboxamide

In a procedure analogous to example 12, reaction of6-chloro-N-[(1S)-1-phenylethyl]pyrazin-2-amine (91 mg, 0.39 mmol) and5-carboxybenzimidazole N-methyl amide (75 mg, 0.43 mmol) furnished thetwo products separated by column chromatography: 5-isomer (23 mg, 16%);6-isomer (38 mg 26%).

(5-isomer) ¹H-n.m.r. (CDCl₃) δ1.67 (d, 3H, J=6.9 Hz, CH₃), 3.06 (d, 3H,J=4.5 Hz, NCH₃), 5.01 (m, 1H, CH), 5.58 (br s, 1H, NH), 6.43 (br s, 1H,CONH), 7.30-7.43 (m, 5H, Ph-H), 7.71 (d, 1H, J=8.4 Hz, benzimid-H),7.84-7.88 (m, 2H, benzimid-H, pyraz-H), 8.19 (s, 1H, pyraz-H), 8.53 (s,1H, benzimid-H), 8.57 (s, 1H, benzimid-H).

m/z (ES) 373 (M⁺+H).

(6-isomer). ¹H-n.m.r. (CDCl₃) δ1.65 (d, 3H, J=6.9 Hz, CH₃), 3.07 (d, 3H,J=4.5 Hz, NCH₃), 5.02 (m, 1H, CH), 5.31 (d, 1H, J=4.8 Hz, NH), 6.30 (brs, 1H, CONH), 7.31-7.41 (m, 5H, Ph-H), 7.68 (d, 1H, J=8.4 Hz,benzimid-H), 7.81 (d, 1H, J=8.4 Hz, benzimid-H), 7.92 (s, 1H, pyraz-H),8.13 (s, 1H, pyraz-H), 8.18 (s, 1H, benzimid-H), 8.49 (s, 1H,benzimid-H).

m/z (ES) 373 (M⁺+H).

EXAMPLE 251-(6-{[(1S)-1-Phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-amineand1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-amine

To a stirred solution of 5-amino-benzimidazole (290 mg, 2.2 mmol) inanhydrous DMF (10 mL) under N₂ was added caesium carbonate (980 mg). Theresulting mixture was stirred at 70 for 60 min. To this was added asolution of 6-chloro-N-[(1S)-1-phenylethyl]pyrazin-2-amine (470 mg) inDMF (5 mL) and the resulting mixture was then heated at reflux for 48 h.The DMF was removed under reduced pressure and the residue diluted withchloroform. The organic layer was washed with aqueous Na₂CO₃, dried(Na₂SO₄) and the solvent removed under reduced pressure to furnish thecrude product. Column chromatography using dichloromethane-methanol(95:5→92:8) as eluant separated two fractions from unreacted startingmaterial. The higher Rf fraction was assigned as the 6-isomer (276 mg,42%). ¹H-n.m.r. (CDCl₃) δ1.64 (d, 3H, J 6.9 Hz, CH₃), 2.90 (br s, 2H,NH₂), 5.05 (m, 1H, CH), 5.21 (d, 1H, NH), 6.70 (dd, 1H, J=8.7, 2.1 Hz,benzimid-H), 6.97 (d, 1H, J=1.8 Hz, benzimid-H), 7.28-7.43 (m, 5H,Ph-H), 7.58 (d, 1H, J=8.4 Hz, benzimid-H), 7.84 (s, 1H, pyraz-H), 8.08(s, H, pyraz-H), 8.21 (s, 1H, benzimid-H). m/z (ES) 331 (M⁺+H). Thelower fraction was assigned as the 5-isomer (170 mg, 26%), ¹H-n.m.r.(CDCl₃) δ1.64 (d, 3H, J=6.9 Hz, CH₃), 2.85 (br s, 2H, NH₂), 5.01 (m, 1H,CH), 5.19 (d, 1H, NH), 6.70 (dd, 1H, 8.7, 2.1 Hz, benzimid-H), 7.11 (d,1H, J=1.8 Hz, benzimid-H), 7.29-7.40 (m, 5H, Ph-H), 7.51 (d, 1H, J=8.7Hz, benzimid-H), 7.81 (s, 1H, pyraz-H), 8.10 (s, 1H, pyraz-H), 8.32 (s,1H, benzimid-H).

m/z (ES) 331 (M⁺+H).

EXAMPLE 26 N-Benzyl-1H-benzimidazole-5-carboxamide

In a procedure analogous to example 20, reaction ofbenzimidazole-5-carboxylic acid (400 mg, 2.50 mmol) and benzylaminefurnished the product (410 mg, 66%).

¹H-n.m.r. (CDCl₃) δ 4.56 (d, 2H, J=5.8 Hz, CH₂), 7.13-7.31 (m, 5H,Ph-H), 7.59 (d, 1H, J=8.5 Hz, H-6), 7.79 (dd, 1H, J=8.5 Hz, H-7), 7.96(br s, 1H, CONH), 8.21 (s, 1H, H-4), 8.29 (s, 1H, H-2).

EXAMPLE 27N-Benzyl-1-[6-([(1S)-1-phenylethyl]amino)pyrazin-2-yl]-1H-benzimidazole-5-carboxamideandN-Benzyl-1-[6-([(1S)-1-phenylethyl]amino)pyrazin-2-yl-1H-benzimidazole-6-carboxamide

In a procedure analogous to example 25, reaction of6-chloro-N-[(1S)-1-phenylethyl]pyrazin-2-amine (91 mg, 0.39 mmol) andN-benzyl-1H-benzimidazole-5-carboxamide (108 mg, 0.43 mmol) furnishedthe product (132 mg, 75%) as a 3:1 mixture of regioisomers.

¹H-n.m.r. (as 3:1 mixture) (CDCl₃) δ1.62 (d, 3H, J=6.9 Hz, CH₃), 1.63(d, 3H, J=CH), 5.55 (d, 1H, NH), 5.61 (d, 1H, NH), 6.80 (t-like, 1H,(CONH), 6.92 (t-like, 1H, CONH), 7.22-7.40 (m, Ph-H, Ph-H), 7.66 (d, 1H,J=8.4 Hz, benzimid-H), 7.71 (d, 1H, J=8.4 Hz, benzimid-H), 7.80-7.85 (m,benzimid-H) 7.91 (s, 1H, benzimid-H), 8.05 (s, 1H, pyraz-H), 8.13 (s,1H, pyraz-H), 8.25 (s, 1H, benzimid-H), 8.38 (s, 1H, pyraz-H), 8.46 (s,1H, benzimid-H), 8.59 (s, 1H, pyraz-H).

m/z (ES) 449 (M⁺+H).

EXAMPLE 28 N-Phenyl-1H-benzimidazole-5-carboxamide

In a procedure analogous to example 20, reaction ofbenzimidazole-5-carboxylic acid (400 mg, 5.50 mmol) and aniline (510 mg,5.50 mmol) furnished the product (370 mg, 63%).

¹H-n.m.r. (CDCl₃) δ6.83 (m, 1H, Ar—H), 7.08 (m, 2H, Ar—H), 7.53-7.74 (m,4H, Ar—H), 8.13-8.31 (m, 2H, Ar—H+CONH), 9.57 (s, 1H, H-2).

EXAMPLE 291-(6-{[(1S)-1-Phenylethyl]amino}pyrazin-2-yl)-N-phenyl-1H-benzimidazole-5-carboxamideand1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-N-phenyl-1H-benzimidazole-6-carboxamide

In a procedure analogous to example 25, reaction of6-chloro-N-[(1S)-1-phenylethyl]pyrazin-2-amine (91 mg, 0.39 mmol) andN-phenyl-1H-benzimidazole-5-carboxamide, (102 mg, 0.43 mmol) furnishedthe product (100 mg, 59%) as a 2:1 mixture of regioisomers.

¹H-n.m.r. (as 2:1 mixture) (CDCl₃) δ1.61 (d, 3H, J=6.9 Hz, CH₃), 1.62(d, 3H, J=6.9 Hz, CH₃), 4.94-5.03 (m, CH, CH), 5.55 (d, 1H, NH), 5.62(d, 1H, NH), 7.18-7.45 (m, Ph-H, Ph-H), 7.61-7.83 (m, benzimid-H andPh-H)), 7.85 (s, 1H, pyraz-H), 7.90 (s, 1H, pyraz-H), 8.06 (s, 1H,pyraz-H), 8.12 (s, 1H, pyraz-H), 8.33 (s, 1H, benzimid-H), 8.36 (s, 1H,benzimid-H), 8.46 (s, 1H, benzimid-H), 8.49 (s, 1H, CONH), 8.52 (s, 1H,CONH), 8.57 (s, 1H, benzimid-H).

m/z (ES) 435 (M⁺+H).

EXAMPLE 30 5-[(4-Methylpiperazin-1-yl)carbonyl]-1H-benzimidazole

In a procedure analogous to example 20, reaction ofbenzimidazole-5-carboxylic acid (400 mg, 2.50 mmol) andN-methylpiperazine (550 mg, 5.0 mmol) furnished the product (380 mg,63%).

¹H-n.m.r. (CDCl₃) δ2.33 (s, 3H, CH₃), 2.44(br s, 4H, CH₂), 3.70 (br s,4H, CH₂), 7.25 (s, 1H, ArH), 7.66 (br s, 1H, ArH), 7.87 (s, 1H, ArH),11.88 (s, 1H, H-2).

m/z (ES) 245 (M⁺+H).

EXAMPLE 316-{5-[(4-Methylpiperazin-1-yl)carbonyl]-1H-benzimidazol-1-yl}-N-[(1S)-1-phenylethyl]pyrazin-2-amineand6-{6-[(4-Methylpiperazin-1-yl)carbonyl]-1H-benzimidazol-1-yl}-N-[(1S)-1-phenylethyl]pyrazin-2-amine

In a procedure analogous to example 25, reaction of6-chloro-N-[(1S)-1-phenylethyl]pyrazin-2-amine (91 mg, 0.39 mmol) and5-[(4-methylpiperazin-1-yl)carbonyl]-1H-benzimidazole (105 mg, 0.43mmol) furnished the product as a mixture of regioisomers which wereseparated by column chromatography: 6-isomer (58 mg, 34%); 5-isomer (68mg, 40%).

(5 isomer) ¹H-n.m.r. (CDCl₃) δ1.63 (d, 3H, J=6.9 Hz, CH₃), 2.33 (s, 3H,NCH₃), 2.44 (br s, 4H, CH₂), 3.67 (br s, 4H, CH₂), 5.01 (m, 1H, CH),5.48 (d, 1H, =6.0 Hz, NH), 7.26-7.38 (m, 6H, Ar—H), 7.65 (d, 1H, J=8.4Hz, benzimid-H), 7.85 (s, 1H, benzimid-H), 7.89 (s, 1H, pyraz-H), 8.09(s, 1H, pyraz-H), 8.41 (s, 1H, benzimid-H).

m/z (ES) 442 (M⁺+H).

(6 isomer) ¹H-n.m.r. (CDCl₃) δ1.63 (d, 3H, J=6.9 Hz, CH₃), 2.31 (s, 3H,NCH₃), 2.43 (br s, 4H, CH₂), 3.4-3.9 (br m, 4H, CH₂), 4.99 (m, 1H, CH),5.54 (d, 1H, J=6.0 Hz, NH), 7.23-7.39 (m, 6H, Ar—H), 7.83-7.85 (m, 2H,Ar—H, pyraz-H), 8.07 (s, 1H, benzimid-H), 8.12 (s, 1H, pyraz-H), 8.44(s, 1H, benzimid-H). m/z (ES) 442 (M⁺+H).

EXAMPLE 32N-[1-(6-{[(1S)-1-Phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]acetamide

To a stirred solution of2-(S-α-methylbenzylamino)-6-(5-amino-benzimidazo-1-yl)-pyrazine (66 mg,0.2 mmol) in anhydrous THF (2 mL) under N₂ was added triethylamine (41mg, 0.4 mmol). The solution was cooled at 0° C. and to this was addedacetyl chloride (17 mg, 0.22 mmol) and the resulting mixture thenstirred at RT. After 18 h the solution was poured into water (30 mL) andthe product extracted into chloroform (2×20 mL). The combined organiclayers were dried (Na₂SO₄) and the solvent removed under reducedpressure to furnish the crude product as a pale yellow solid. Columnchromatography using dichloromethane-methanol (200:15) as eluantseparated the product as a pale yellow solid (38 mg).

¹H-n.m.r. (CDCl₃) δ1.63 (d, 3H, J=6.6 Hz, CH₃), 2.21 (s, 3H, CH₃), 5.00(m, 1H, CH), 5.43 (d, 1H, J=5.7 Hz, NH), 7.27-7.38 (m, 5H, ArH), 7.49(d, 1H, J=9.0 Hz, benzimid-H), 7.61 (d, 1H, J=9.0 Hz, benzimid-H), 7.74(br s, 1H, CONH), 7.84 (s, 1H, pyraz-H), 7.90 (s, 1H, benzimid-H), 8.11(s, 1H, pyraz-H), 8.36 (s, 1H, benzimid-H).

m/z (ES) 373. (M⁺+H).

EXAMPLE 33N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]acetamide

In a procedure analogous to example 32, reaction of2-(S-α-methylbenzylamino)-6-(6-amino-benzimidazo-1-yl)-pyrazine (66 mg,0.20 mmol) and acetyl chloride (17 mg, 0.22 mmol) furnished the product(70 mg, 94%) after chromatography.

¹H-n.m.r. (CDCl₃) δ1.65 (d, 3H, J=6.9 Hz, CH₃), 2.22 (s, 3H, CH₃), 5.07(m, 1H, CH), 5.29 (d, 1H, J=6.3 Hz, NH), 7.28-7.43 (m, 6H, ArH,benzimid-H), 7.72 (d, 1H, J=8.7 Hz, benzimid-H), 7.84 (s, 1H, pyraz-H)8.12 (s, 1H, pyraz-H), 8.27 (s, 1H, benzimid-H), 8.34 (s, 1H,benzimid-H).

m/z (ES) 373 (M⁺+H).

EXAMPLE 34N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]benzamide

In a procedure analogous to example 32, reaction of1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-amine(33 mg, 0.10 mmol) and benzoyl chloride (14 mg 0.10 mmol) furnished theproduct (23 mg, 53%) after chromatography.

¹H-n.m.r. (CDCl₃) δ 1.64 (d, 3H, J=6.8 Hz, CH₃), 5.02 (m, 1H, CH), 5.33(d, 1H, J=4.5 Hz, NH), 7.27-7.68 (m, 10H, ArH) 7.85 (s, 1H, pyraz-H),7.90-7.93 (m, 2H, benzimid-H), 8.03 (s, 1H, benzimid-H), 8.06 (s 1H,CONH), 8.10 (s, 1H, pyraz-H), 8.38 (s, 1H, benzimid-H).

m/z (ES) 435 (M⁺+H).

EXAMPLE 35N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]benzamide

In a procedure analogous to example 32, reaction of1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-amine(50 mg, 0.15 mmol) and benzoyl chloride (21 mg, 0.15 mmol) furnished theproduct (50 mg, 76%) after chromatography.

¹H-n.m.r. (CDCl₃) δ1.64 (d, 3H, J=6.6 Hz, CH₃), 5.11 (m, 1H, CH), 5.31(d, 1H, J=6.6 Hz, NH), 7.16-7.58 (m, 10H, ArH), 7.78 (d, 1H, J=8.4 Hz,benzimid-H), 7.83 (s, 1H, pyraz-H), 8.35 (s, 1H, pyraz-H), 8.51 (s, 1H,benzimid-H).

m/z (ES) 435 (M⁺+H).

EXAMPLE 36N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]isonicotinamide

In a procedure analogous to example 32, reaction of1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-amine(33 mg, 0.10 mmol) and isonicotinoyl chloride hydrochloride (20 mg, 0.11mmol) furnished the product (10 mg, 23%) after chromatography.

¹H-n.m.r. (CDCl₃) δ1.56 (d, 3H, J=6.9 Hz, CH₃), 4.98 (m, 1H, CH), 6.21(d, 1H, J=6.0 Hz, NH), 7.11-7.37 (m, 5H, ArH), 7.59 (d, 1H, J=9.0 Hz,benzimid-H), 7.65 (d, 1H, J=8.7 Hz, benzimid-H), 7.81 (d, 2H, J=5.7 Hz,pyrid-H), 7.87 (s, 1H, pyraz-H), 8.01 (s, 1H, pyraz-H), 8.10 (s, 1H,benzimid-H), 8.32 (s, 1H, benzimid-H), 8.72 (d, 2H, J=5.7 Hz, pyrid-H),9.58 (s, 1H, CONH).

m/z (ES) 436 (M⁺+H).

EXAMPLE 37N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]isonicotinamide

In a procedure analogous to example 32, reaction of1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-amine(33 mg, 0.10 mmol) and isonicotinoyl chloride hydrochloride (20 mg, 0.11mmol) furnished the product (10 mg, 23%) after chromatography.

¹H-n.m.r. (CDCl₃) δ1.63 (d, 3H, J=6.9 Hz, CH₃), 5.11 (m, 1H, CH), 5.40(d, 1H, J=6.3 Hz, NH), 7.18-7.42 (m, 6H, ArH, CONH), 7.73-7.79 (m, 3H,benzimid-H), 7.83 (s, 1H, pyraz-H), 8.10 (s, 1H, pyraz-H), 8.36 (br s,2H, pyrid-H), 8.48 (s, 1H, benzimid-H), 8.79 (d, 2H, J=5.7 Hz, pyrid-H).

m/z (ES) 436 (M⁺+H).

EXAMPLE 38N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5yl]nicotinamide

In a procedure analogous to example 32, reaction of1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-amine(33 mg, 0.10 mmol) and nicotinoyl chloride hydrochloride (20 mg, 0.11mmol) furnished the product (17 mg, 39%) after chromatography.

¹H-n.m.r. (CDCl₃) δ1.48 (d, 3H, J=6.9 Hz, CH₃), 4.91 (m, 1H, CH), 6.59(d, 1H, J=5.7 Hz, NH), 7.11-7.32 (m, 6H, ArH, pyrid-H), 7.58 (d, 1H,J=9.0 Hz, benzimid-H), 7.83 (s, 1H, benzimid-H), 7.91 (s, 1H, pyraz-H),8.07 (s, 1H, benzimid-H), 8.20(d, 1H, J=8.1 Hz, pyrid-H), 8.24 (s, 1H,pyraz-H), 8.61 (d, 1H, J=4.6 Hz, pyrid-H), 9.12 (s, 1H, pyrid-H), 9.80(s, 1H, CONH).

m/z (ES) 436 (M⁺+H).

EXAMPLE 39N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]nicotinamide

In a procedure analogous to example 32, reaction of1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-amine(33 mg, 0.10 mmol) and isonicotinoyl chloride hydrochloride (20 mg, 0.11mmol) furnished the product (27 mg, 62%) after chromatography.

¹H-n.m.r. (CDCl₃) δ1.62 (d, 3H, J=6.6 Hz, CH₃), 5.12 (m, 1H, CH), 5.41(d, 1H, J=6.0 Hz, NH), 7.17-7.21 (m, 1H, Ar—H), 7.26-7.29 (m, 1H, Ar—H),7.39-7.45 (m, 4H, ArH, pyrid-H), 7.76 (d, 1H, J=8.7 Hz, benzimid-H),7.82 (s, 1H, pyraz-H), 8.11 (s, 1H, pyraz-H), 8.21 (d, 1H, J=8.1 Hz,pyrid-H), 8.33 (s, 1H, CONH). 8.35 (s, 1H, Ar—H), 8.47 (3, 1H, pyrid-H),8.76 (1H, d, J=4.5 Hz, pyrid-H), 9.13 (s, 1H, pyrid.-H).

m/z (E) 436 (M⁺+H).

EXAMPLE 40N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]pyrazine-2-carboxamide

In a procedure analogous to example 32, reaction of1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-amine(33 mg, 0.10 mmol) and pyrazine-2-carbonyl chloride (0.11 mmol)(prepared by reacting pyrazine-2-carboxylic acid with oxalyl chloride indicloromethane at room temperature) furnished the product (24 mg 55%)after chromatography.

¹H-n.m.r. (CDCl₃) δ1.65 (d, 3H, J=6.9 Hz, CH₃), 5.04 (m, 1H, CH), 5.35(d, 1H, J=6.0 Hz, NH), 7.28-7.41 (m, 5H, Ar—H, pyrid-H), 7.71 (s, 1H,pyraz-H), 7.87 (s, 1H, pyraz-H), 8.12 (s, 1H, pyraz-H), 8.21 (s, 1H,benzimid-H), 8.40 (s, 1H, pyraz-H), 8.61-8.62 (m, 1H, benzimid-H), 8.81(d, 1H, J=2.4 Hz, benzimid-H), 9.55 (d, 1H, J=1.2 Hz, benzimid-H), 9.78(s, 1H, CONH).

m/z (ES) 437 (M⁺+H).

EXAMPLE 41N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]pyrazine-2-carboxamide

In a procedure analogous to example 32, reaction of1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-amine(33 mg, 0.10 mmol) and isonicotinoyl chloride hydrochloride (0.11 mmol)(prepared by reacting pyrazine-2-carboxylic acid with oxalyl chloride indicloromethane at room temperature) furnished the product (28 mg, 64%)after chromatography.

¹H-n.m.r. (CDCl₃) δ1.67 (d, 3H, J=6.9 Hz, CH₃), 5.18 (m, 1H, CH), 5.39(d, 1H, J=6.3 Hz, NH), 7.23-7.46 (m, 6H, Ar—H, pyrid-H), 7.82 (d, 1H,J=9.0 Hz, benzimid-H), 7.84 (s, 1H, pyraz-H), 8.18 (s, 1H, pyraz-H),8.39 (s, 1H, pyraz-H), 8.61-8.62 (m, 1H, pyraz-H), 8.81-8.84 (m, 2H,—Ar—H), 9.52 (d, 1H, J=1.2 Hz, benzimid-H), 9.81 (s, 1H, CONH).

m/z (ES) 437 (M⁺+H).

EXAMPLE 422-Methoxy-N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]acetamide

In a procedure analogous to example 32, reaction of1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-amine(33 mg, 0.10 mmol) and methoxyacetyl chloride (12 mg, 0.11 mmol)furnished the product (20 mg, 50%) after chromatography.

¹H-n.m.r. (CDCl₃) δ1.64 (d, 3H, J=6.9 Hz, CH₃), 3.55 (s, 3H, OCH₃), 4.07(s, 2H, CH₂), 5.02 (m, 1H, CH), 5.27 (d, 1H, J=6.3 Hz, NH), 7.30-7.40(m, 5H, Ar—H), 7.54 (dd, 1H, J=8.7, 1.5 Hz, benzimid-H), 7.66 (d, 1H,J=8.7 Hz, benzimid-H), 7.85 (s, 1H, pyraz-H), 8.01 (s, 1H, benzimid-H),8.11 (s, 1H, pyraz-H), 8.35 (s, 1H, CONH), 8.38 (s, 1H, benzimid-H).

m/z (ES) 403 (M⁺+H).

EXAMPLE 432-Methoxy-N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]acetamide

In a procedure analogous to example 32, reaction of1-(6-{[91S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-amine(33 mg, 0.10 mmol) and methoxyacetyl chloride (20 mg, 0.11 mmol)furnished the product (10 mg, 25%) after chromatography.

¹H-n.m.r. (CDCl₃) δ1.64 (d, 3H, J=6.9 Hz, CH₃), 3.54 (s, 3H, OCH₃), 4.06(s, 2H, CH₂), 5.08 (m, 1H, CH), 5.36 (d, 1H, 6.3 Hz, NH), 7.24-7.42 (m,6H, Ar—H), 7.76 (d, 1H, J=8.7 Hz, benzimid-H), 7.81 (s, 1H, pyraz-H),8.14 (s, 1H, pyraz-H), 8.35 (s, 1H, benzimid-H), 8.39 (s, 1H, CONH),8.52 (d, 1H, 1.5 Hz, benzimid-H).

m/z (ES) 403 (M⁺+H).

EXAMPLE 44N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-6-yl]-2,2-dimethylpropanamide

To a stirred solution of2-(benzylamino)-6-(5-amino-benzimidazo-1-yl)-pyrazine (33 mg, 0.1 mmol)in anhydrous THF (2 mL) under N₂ was added triethylamine (38 l, 0.3mmol). The solution was cooled at 0° C. and to this was added pivalicacid (12 mg, 0.11 mmol) and EDC (23 mg, 0.12 mmol) and the resultingmixture then stirred at RT. After 64 h the solution was diluted with H₂Oand the mixture extracted with CHCl₃ (2×15 mL). The combined organiclayers were washed with 10% aqueous Na₂CO₃, dried (Na₂SO₄) and thesolvent removed in vacuo. The residue was purified by columnchromatography using dichloromethane-methanol (100:6) as eluant toseparate the pur product (15 mg).

¹H-n.m.r. (CDCl₃) δ1.35 (s, 9H, 3CH₃), 1.65 (d, 3H, J=6:6 Hz, CH₃), 5.14(m, 1H, CH), 5.24 (d, 1H, J=5.7 Hz, NH), 7.13 (d, 1H, J=8.7 Hz, Ar—H),7.29-7.47 (m, 5H, ArH), 7.75 (d, 1H, J=8.7 Hz, benzimid-H), 7.81 (s, 1H,pyraz-H), 8.17 (s, 1H, pyraz-H), 8.35 (s, 1H, benzimid-H), 8.69 (s, 1H,CONH).

m/z (ES) 415 (M⁺+H).

EXAMPLE 45N-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]-2,2-dimethylpropanamide

In a procedure analogous to example 44, reaction of1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-amine(33 mg, 0.10 mmol) and pivalic acid (12 mg, 0.11 mmol) furnished theproduct (12 mg, 29%) after chromatography.

¹H-n.m.r. (CDCl₃) δ1-35 (s, 9H, CH₃), 1.66 (d, 3H, J=6.9 Hz, CH₃), 5.14(m, 1H, CH), 5.24 (d, 1H, J=6.3 Hz, NH), 7.13 (d, 1H, J=8.7 Hz, Ar—H),7.29-7.47 (m, 6H, ArH), 7.75 (d, 1H, J=8.7 Hz, benzimid-H), 7.81 (s, 1H,pyraz-H), 8.17 (s, 1H, pyraz-H), 8.35 (s, 1H, benzimid-H), 8.69 (s, 1H,CONH).

m/z (ES) 415 (M⁺+H).

EXAMPLE 466-{5-[(4-Methylpiperazin-1-yl)methyl]-1H-benzimidazol-1-yl}-N-[(1S)-1-phenylethyl]pyrazin-2-amine

A solution of6-{5-[(4-methylpiperazin-1-yl)carbonyl]-1H-benzmidazol-1-yl}-N-[(1S)-1-phenylethyl]pyrazin-2-amine(22 mg, 0.05 mmol) in dry THF (1 mL) was added to a suspension of LiAlH₄(4 mg, 0.1 mmol) in THF (1 mL) and the mixture heated at reflux for 4 h.Upon cooling to RT, the solution was treated consecutively with H₂O (1mL), aqueous NaOH (1 mL, 2M) and H₂O (5 mL). The resulting mixture wasextracted with CHCl₃ (2×10 mL) and the combined organic layers dried(Na₂SO₄). The solvent was removed under reduced pressure and the productpurified by flash chromatography using CH₂Cl₂—MeOH (10:1→1:1) as eluantto afford the product as a yellow solid (11 mg, 52%).

¹H-n.m.r. (CDCl₃) δ1.65 (d, 3H, J=6.9 Hz, CH₃), 2.58 (s, 3H, NCH₃), 2.81(br s, 4H, CH₂), 2.90 (br s, 4H, CH₂), 3.74 (s, 2H, NCH₂), 5.03 (m, 1H,CH), 5.33 (d, 1H, J=6.0 Hz, NH), 7.25-7.42 (m, 6H, Ar—H), 7.67 (d, 1H,J=8.4 Hz, benzimid-H), 7.77 (s, 1H, benzimid-H), 7.87 (s, 1H, pyraz-H),8.12 (s, 1H, pyraz-H), 8.39 (s, 1H, benzimid-H).

m/z (ES) 428 (M⁺+H).

EXAMPLE 471-[6-{[(1S)-1-Phethylethyl]amino}pyrazin-2-yl]-N-(pyridin-3-ylmethyl)-1H-benzimidazol-5-amine

In a procedure analogous to example 46, reaction ofN-[1-(6-{[(1S)-1-phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl]nicotinamide(33 mg, 0.10 mmol) with LiAlH₄ (5.7 mg, 0.15 mmol) furnished the product(10 mg, 24%) after chromatography.

¹H-n.m.r. (CDCl₃) δ 1.64 (d, J=6.9 Hz, 3H, CH₃), 4.42(s, 2H, CH₂),5.01(m, 1H, CH), 5.20(d, J=6.0 Hz, 1H, NH), 6.66 (dd, J=8.7, 2.1 Hz, 1H,H-7′), 7.01 (d, J=2.1 Hz, 1H, H-4′), 7.25-7.45 (m, 6H, ArH), 7.51 (d,J=8.7 Hz, 1H, H-6′), 7.74 (d, J=7.8 Hz, pyridine-H), 7.80 (s, 1H,pyraz-H), 8.09 (s, 1H, pyraz-H), 8.30 (s, 1H, H-2′), 8.53 (d, J=3.6 Hz,1H, pyridine-H), 8.68(s, 1H, pyridine-H).

m/z (ES) 422 (M⁺+H).

EXAMPLE 48 N-[(1S)-1-(4-Bromophenyl)ethyl]-6-chloropyrazin-2-amine

In a procedure analogous to example 1, reaction of4-bromo-α-S-methyl-benzylamine (877 mg, 4.4 mmol) and2,6-dichloropyrazine (597 mg, 4.0 mmol) furnished the product (835 mg,67%).

¹H-n.m.r. (CDCl₃) δ 1.56 (d, J=6.9 Hz, 3H, CH₃), 4.86 (m, 1H, CH), 5.0(d, 1H, NH), 7.24 (AA′XX′, 2H, ArH) 7.60 (s, 1H, pyraz-H), 7.81 (s, 1H,pyraz-H).

EXAMPLE 49 6-Chloro-N-[(1S)-1-(4-methoxyphenyl)ethyl]pyrazin-2-amine

In a procedure analogous to examples 1, reaction of4-methoxy-α-S-methyl-benzylamine (700 mg, 4.6 mmol) and2,6-dichloropyrazine (626 mg, 4.2 mmol) furnished the product (873 mg,79%).

¹H-n.m.r. (CDCl₃) δ1.56 (d, J=6.9 Hz, 3H, CH₃), 3.80 (s, 3H, OCH₃), 4.84(m, 1H, CH), 5.01 (d, 1H, NH), 6.88 (AA′XX′, 2H, Ar—H), 7.28 (AA′XX′,2H, Ar—H), 7.621 (s, 1H, pyraz-H), 7.79 (s, 1H, pyraz-H).

EXAMPLE 506-(1H-Benzimidazol-1-yl)-N-[(1S)-1-(4-bromophenyl)ethyl]pyrazin-2-amine

In a procedure analogous to example 25, reaction ofN-[(1S)-1-(4-bromophenyl)ethyl]-6-chloropyrazin-2-amine (125 mg, 0.40mmol) and benzimidazole (52 mg, 0.44 mmol) furnished the product (66 mg,42%).

¹H-n.m.r. (CDCl₃) δ1.63 (d, 3H, J=6.9 Hz, CH₃), 4.99 (m, 1H, CH), 5.19(d, 1H, J=5.1 Hz, NH), 7.26-7.37 (m, 3H, Ar—H), 7.51 (AA′XX′, 2H, Ar—H),7.65 (d, 1H, J=8.1 Hz, benzimid-H), 7.83-7.86 (m, 2H,benzimid-H+pyraz-H), 8.17 (s, 1H, pyraz-H), 8.39 (s, 1H, benzimid-H).

m/z (ES) 396, 394 (M⁺+H)

EXAMPLE 516-(1H-Benzimidazol-1-yl)-N-[(1S)-1-(4-methoxyphenyl)ethyl]pyrazin-2-amine

In a procedure analogous to example 25, reaction of6-chloro-N-[(1S)—1-(4-methoxyphenyl)ethyl]pyrazin-2-amine (105 mg, 0.40mmol) and benzimidazole (52 mg, 0.44 mmol) furnished the product (57 mg,41%).

¹H-n.m.r. (CDCl₃) δ1.62 (d, 3H, J=6.8 Hz, CH₃), 3.80 (s, 3H, CH₃), 4.99(m, 1H, CH), 5.31 (br d, 1H, J=6.2 Hz, NH), 6.91 (AA′XX′, 2H, Ar—H),7.28-7.36 (m, 4H, Ar—H), 7.77 (d, 1H, J=2.0 Hz, Ar—H), 7.78 (s, 2H,benzimid-H+pyraz-H), 8.13 (s, 1H, pyraz-H), 8.44 (s, 1H, benzimid-H).

m/z (ES) 346 (M⁺+H)

EXAMPLE 522-(S-α-Methylbenzylamino)-6-(5-(N-methylpiperazin-4-yl-methyl)-benzimidazo-1-yl)-pyrazine

A solution of3-[6-(S-α-methylbenzylamino)-pyrazin-2-yl]-3H-benzoimidazole-5-carboxylicacid N-methylpiperazinylamide (22 mg, 0.05 mmol) in dry THF (1 mL) wasadded to a suspension of LiAlH₄ (4 mg, 0.1 mmol) in THF (1 mL) and themixture heated at reflux for 4 h. Upon cooling to RT, the solution wastreated consecutively with H₂O (1 mL), aqueous NaOH (1 mL, 2M) and H₂O(5 mL). The resulting mixture was extracted with CHCl₃ (2×10 mL) and thecombined organic layers dried (Na₂SO₄). The solvent was removed underreduced pressure and the product purified by flash chromatography usingCH₂Cl₂-MeOH (10:1→1:1) as eluant to afford the product as a yellow solid(11 mg, 52%).

¹H-n.m.r. (CDCl₃) δ1.65 (d, 3H, J=6.9 Hz, CH₃), 2.58 (s, 3H, NCH₃), 2.81(br s, 4H, CH₂), 2.90 (br s, 4H, CH₂), 3.74 (s, 2H, NCH₂), 5.03 (m, 1H,CH), 5.33 (d, 1H, J=6.0 Hz, NH), 7.25-7.42 (m, 6H, Ar—H), 7.67 (d, 1H,J=8.4 Hz, benzimid-H), 7.77 (s, 1H, benzimid-H), 7.87 (s, 1H, pyraz-H),8.12 (s, 1H, pyraz-H), 8.39 (s, 1H, benzimid-H).

m/z (ES) 428 (M⁺+H).

EXAMPLE 53 1-{4-[2-(Diethylamino)ethoxy]phenyl}ethanone

A mixture of 4-bromoacetophenone (5 g, 25 mmol), the diethylaminoethanol(3.5 g, 38 mmol), K₂CO₃ (2g), copper powder (0.5 g) and copper(I) iodide(2.5 g) in DMSO (30 ml) was heated at 120° C. until TLC showedconsumption of the starting material. After cooling to RT, the mixturewas poured onto aqueous NH₃ (28%, 100 ml), extracted with CH₂Cl₂ (3×100ml). The combined organic extracts were dried (Na₂SO₄), filtered and theCH₂Cl₂ distilled in vacuo. The crude product was purified by columnchromatography on silica gel, eluting with EtOAc-MeOH (95:5) to furnishthe pure compound (4.85 g, 82%).

¹H-n.m.r. (CDCl₃) δ0.98 (t, J=7.2 Hz, 6H, NCH₂CH₃), 2.43 (s, 3H,Ar—C—CH₃), 2.57-2.50 (m, 4H, NCH₂CH₃), 2.78 (t, J=6.3 Hz, 2H,ArOCH₂CH₂N), 4.00 (t, J=6.3 Hz, 2H, ArO-CH₂), 6.85 (d, J=9 Hz, 2H, ArH),7.83 (d, J=9 Hz, 2H, ArH).

EXAMPLE 54 1-[4-(4-Methylpiperazin-1-yl)phenyl]ethanone

In a procedure identical to Example 53, 4-bromoacetophenone was reactedwith N-methylpiperazine in the presence of potassium carbonate, copperand copper iodide to furnish the desired product in 82% yield.

¹H-n.m.r. (CDCl₃) δ 2.35 (s, 3H, Ar—C—CH₃), 2.52 (s, 3H, N—CH₃), 2.55(t, J=5.1 Hz, 4H, CH₂—N—CH₃), 3.37 (dd, J=5.1, 5.1 Hz, 4H, ArNCH₂), 6.89(AA′XX′, 2H, ArH), 7.89 (AA′XX′, 2H, ArH).

EXAMPLE 55 1-Pyridin-3-ylethanone oxime

To a solution of hydroxylamine hydrochloride (3.44 g) in water (20 ml)was added NaOH (20%, 30 ml). The ketone (5 g, 41 mmol) was added at onceand the resulting mixture was stirred at RT until TLC showed no ketoneremained. The solvents were distilled off in vacuo and the residueextracted with CH₂Cl₂ (3×100 ml) and dried (Na₂SO₄). After filtrationand removal of the solvent, the crude ketoxime was recrystallised fromCH₂Cl₂/n-hexane.

¹H-n.m.r. (CDCl₃) δ2.31 (s, 3H, CH₃), 7.33 (dd, J=4.8, 4.8 Hz, 1H, ArH),7.97 (ddd, J=8.1, 1.8, 1.8 Hz, 1H, ArH), 8.61 (dd, J=5.1, 1.8 Hz, 1H,ArH), 8.96 (d, J=1.8 Hz, 1H, ArH), 10.62 (s, 1H, OH).

EXAMPLE 56 1-(3-Chlorophenyl)ethanone oxime

A mixture of the ketone (2.0 g, 13 mmol), hydroxylamine hydrochloride(0.98 g, 14 mmol), NaOH (10%, 4 ml), water (6.2 ml) and EtOH (25 ml) washeated under reflux for 2 hours. Upon cooling in ice, the ketoximeprecipitated and was collected by suction filtration. The crude productwas recrystallised from CH₂C₂/n-hexane (1.88 g, 86%).

¹H-n.m.r. (CDCl₃) δ 2.28 (s, 3H, CH₃), 7.51 (s, 4H, ArH), 8.67 (s, 1H,OH).

EXAMPLE 57 1-(3-Chlorophenyl)ethanamine

A mixture of the ketoxime (1 g, 6 mmol) and LiAlH₄ (0.27 g) in anhydrousTHF (100 ml) was heated at reflux under dry N₂ overnight. The reactionmixture was cooled in ice-water and carefully quenched with H₂O (60 mL).The mixture was allowed to stir at RT for half an hour, after which timeit was filtered through Celite®. The inorganic salts were washed withEtOAc (3×100 ml). The filtrate was concentrated under reduced pressure,diluted with 2M HCl (50 ml) and the aqueous phase washed with Et₂O (2×70ml). The aqueous phase was basified with 40% aqueous NaOH and theproduct extracted with Et₂O (3×50 ml). The combined organic layers werewashed with brine (50 ml) and dried (MgSO₄). The solvents were removedin vacuo to afford the pure amine (0.65 g, 71%).

¹H-n.m.r. (CDCl₃) δ1.38 (d, J=6.6 Hz, 3H, CH—CH₃), 1.63 (br s, 2H, NH₂),4.13-4.06 (m, 1H, CH—CH₃), 7.23-7.18 (m, 3H, ArH), 7.35 (s, 1H, ArH).

EXAMPLE 58 1-Pyridin-3-ylethanamine

To a mixture of the ketoxime (4.85 g, 36 mmol) and Zn powder (12 g) at0° C. was slowly added, with vigorous stirring, concentrated HCl (50ml). When the initial vigorous reaction had subsided, the mixture washeated under reflux until TLC showed all the ketoxime had been consumed.After cooling to RT, the strongly acidic mixture was extracted withCH₂Cl₂ (2×75 ml). The reaction mixture was then made strongly basic with50% KOH solution. After removal of the solvent, the residue wasextracted with boiling MeOH (4×100 ml). The MeOH was distilled off toleave the crude amine which was used in the ensuing reactions withoutfurther purification.

¹H-n.m.r. (CDCl₃) δ1.07 (d, J=6.6 Hz, 3H, CH₃), 1.37 (br s, 2H, NH₂),3.84 (q, J=4.6 Hz, 1H, CH—CH₃), 6.93 (dd, J=7.8, 4.8 Hz, 1H, ArH), 7.38(ddd, J=7.8, 2.1, 1.5 Hz, 1H, ArH), 8.15 (dd, J=4.8, 1.5 Hz, 1H, ArH),8.27 (d, J=2.1 Hz, 1H, ArH).

EXAMPLE 596-Chloro-N-[(1S)-1-(4-pyridin-3-ylphenyl)ethyl]pyrazin-2-amine

Under a nitrogen atmosphere a mixture ofN-[(1S)-1-(4-bromophenyl)ethyl]-6-chloropyrazin-2-amine (0.117 g, 0.37mmol), pyridine-3-boronic acid 1,3-propanediol cyclic ester (67 mg, 0.41mmol), tetrakis(triphenylphosphine)palladium(0) (65 mg, 0.06 mmol) andtoluene (4 mL) was treated with 2M aqueous sodium carbonate solution(0.2 mL). The resulting mixture was stirred vigorously whilst beingheated under reflux for 24 hours. Upon cooling, the solution was dilutedwith methanol and dichloromethane and the mixture dried (MgSO₄) andfiltered. Removal of solvent in vacuo then yielded the crude productwhich was purified by column chromatography usingdichloromethane-diethyl ether (90:10) then dichloromethane-methanol(99:1) as eluent (50 mg).

¹H-n.m.r. (CDCl₃) δ1.61 (d, 3H, J=6.9 Hz, CH₃), 4.97 (m, 1H, CH), 5.42(d, 1H, J=6.3 Hz, NH), 7.33-7.37 (m, 1H, ArH), 7.42-7.56 (m, 4H, ArH),7.66 (m, 1H, pyraz-H), 7.78 (s, 1H, pyraz-H), 7.83-7.86 (m, 1H, ArH),8.58 (br s, 1H, pyrid-H), 8.83 (br s, 1H, pyrid-H).

m/z (ES) 313, 311 (M++H).

EXAMPLE 60N-[1-(6-{[(1S)-1-Phenylethyl]amino}pyrazin-2-yl)-1H-benzimidazol-5-yl)]methanesulfonamide

To a stirred solution of2-(S-α-methylbenzylamino)-6-(5-amino-benzimidazo-1-yl)-pyrazine (33 mg,0.1 mmol) in anhydrous THF (2 mL) under N₂ was added triethylamine (40mg, 0.4 mmol). The solution was cooled at 0° C. and to this was addedmethanesulphonyl chloride (25 mg, 0.2 mmol) and the resulting mixturethen stirred at RT. After 16 h the solution was poured into water (30mL) and the product extracted into chloroform (2×15 mL). The combinedorganic layers were was washed with 10% Na₂CO₃, dried (Na₂SO₄) and thesolvent removed under reduced pressure to furnish the crude product as apale yellow solid. Column chromatography, using dichloromethane-methanol(100:6) as eluant, separated the product from the most polar fractionsas a pale yellow solid (16 mg).

¹H-n.m.r. (CDCl₃) δ1.65 (d, 3H, J=6.9 Hz, CH₃), 3.00 (s, 3H, CH₃), 5.02(m, 1H, CH), 5.27 (d, H, J=6.0 Hz, NH), 7.21-7.40 (m, 6H, ArH), 7.64 (d,1H, J=8.7 Hz, benzimid-H), 7.69 (d, 1H, J=1.9 Hz, benzimid-H), 7.88 (s,1H, pyraz-H), 8.10 (s, 1-H, pyraz-H), 8.41 (s, 1H, benzimid-H).

m/z (ES) 409 (M++H).

EXAMPLE 61N-[(1S)-1-Phenylethyl]-6-(5-pyridin-4-yl-1H-benzimidazol-1-yl)pyrazin-2-amineandN-[(1S)-1-Phenylethyl]-6-(6-pyridin-4-yl-1H-benzimidazol-1-yl)pyrazin-2-amine

To a stirred solution of a 1:1 mixture of6-(5-bromo-1H-benzimidazol-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amineand6-(6-bromo-1H-benzimidazol-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine(16 mg, 0.4 mmol), pyridine-4-boronic acid pinacol cyclic ester (91 mg,0.44 mmol) in toluene (5 mL) was added tetrakis(triphenyl-phosphine)palladium (46 mg, 0.04 mmol) followed by aqueous sodium carbonate (0.22mL, 2M). The solution was then heated under reflux for 24 h. Afterstandard work-up (see example 60) the products were obtained, as a 1:1mixture by flash chromatography using dichloromethane-methanol (100:6)as eluant.

¹H-n.m.r. (as 1:1 mixture) (CDCl₃) δ 1.66 (d, J=6.9 Hz, 3H, CH₃), 5.07(m, 1H, CH), 5.25 (d, J=6.0 Hz, 1H, NH), 7.29-7.42 (m, 5H, ArH),7.52-7.64 (m, 3H), 7.78 (d, J=8.7 Hz), 7.88 and 7.90 (s, 1H, pyraz-H),7.94 (d, 1H, J=8.4 Hz) 8.12 (s, 1H), 8.16 (s, 1H), 8.19 and 8.20 (s, 1H,pyraz-H), 8.44 and 8.45 (s, 1H, H-2′), 8.68 (d, J=3.6 Hz, 1H,pyridine-H), 8.68 and 8.71 (d, 1H, J=8.7 Hz, pyridine-H).

m/z (ES) 393 (M⁺+H).

Screening

Compound Dilution

For screening purposes, compounds were diluted in 96 well plates at aconcentration of 20 μM. Plates were warmed at 37° C. for 30 minutesbefore assay.

JAK Tyrosine Kinase Domain Production

JAK kinase domains were produced in the following manner:

JAK1

The kinase domain of humanJAK1 was amplified from U937mRNA using thepolymerase chain reaction with the following primers:

XHOI-J1 (SEQ ID NO:1) 5′-CCG CTC GAG ACT GAA GTG GAC CCC ACA CAT-3′J1-KPNI (SEQ ID NO:2) 5′-CGG GGT ACC TTA TTT TAA AAG TGC TTC AAA-3′JAK1 PCR products were cloned into the pFastBac HTb expression vector(Gibco) via the Xho I and Kpn I sites. The JAK1 plasmid was thentransformed into competent DH10Bac cells (Gibco), and the recombinantbaculovirus produced prepared for transfection into Sf9 insect cells.JAK2The kinase domain of humanJAK2 was amplified from U937mRNA using thepolymerase chain reaction with the following primers:

SALI-jk2 (SEQ ID NO:3) 5′-ACG CGT CGA CGG TGC CTT TGA AGA CCG GGA T-3′jk2-NOTI 5′- (SEQ ID NO:4) ATA GTT TAG CGG CCG CTC AGA ATG AAG GTC ATTT-3′JAK2 PCR products were cloned into the pFastBac HTc expression vector(Gibco) via the Sal I and Not I sites. The JAK2 plasmid was thentransformed into competent DH10Bac cells (Gibco), and the recombinantbaculovirus produced prepared for transfection into Sf9 insect cells.JAK3The kinase domain of humanJAK3 was amplified from U937mRNA using thepolymerase chain reaction with the following primers:

XHOI-J3 (SEQ ID NO:5) 5′-CCG CTC GAG TAT GCC TGC CAA GAC CCC ACG-3′J3-KPNI (SEQ ID NO:6) 5′-CGG GGT ACC CTA TGA AAA GGA CAG GGA GTG-3′JAK3 PCR products were cloned into the pFastBac HTb expression vector(Gibco) via the Xho I and Kpn I sites. The JAK3 plasmid was thentransformed into competent DH10Bac cells (Gibco), and the recombinantbaculovirus produced prepared for transfection into Sf9 insect cells.TYK2The kinase domain of humanTYK2 was amplified from A549 mRNA using thepolymerase chain reaction with the following primers:

HT2EK (SEQ ID NO:7) 5′-GGA GCA CTC GAG ATG GTA GCA CAC AAC CAG GTG-3′ITY2.2R 5′- (SEQ ID NO:8) GGA GCA GGA ATT CCG GCG CTG CCG GTC AAA TCTGG-3′TYK2 PCR products were cloned into pBlueBacHis2A (Invitrogen) via theEcoRI site. The recombinant TYK2 baculovirus produced was prepared fortransfected into Sf9 insect cells.Large Scale Production Of Kinase Domains

Baculovirus preparations from each of the JAK family members wereinfected into five litres of High Five cells (Invitrogen) grown in HighFive serum free medium (Invitrogen) to a cell density of approximately1-2×10⁶ cells/ml. Cells are infected with virus at a MOI of 0.8-3.0.Cells were harvested and lysed. JAK kinase domains were purified byaffinity chromatography on a Probond (Invitrogen) nickel chelateaffinity column.

Assay Protocols

Kinase assays were performed either in a 96 well capture-based ELISAassay or in 384 well Optiplates (Packard) using an Alphascreen ProteinTyrosine Kinase kit. In either case using approximately 1.5 μg ofaffinity purified PTK domain in the presence of 50 mM HEPES, pH 7.5, 10mM MgCl_(2, 150) mM NaCl and 10 μM-1mM ATP. The biotinylated substratebiotin-EGPWLEEEEEAYGWMDF-NH₂ (SEQ ID NO:9) (final concentration 5 μM)was used as substrate. In the ELISA assay tyrosine phosphorylation wasquantitated following transfer to an avidin coated ELISA plate usingperoxidase-linked anti-phospho-tyrosine antibody PY20. In theAlphascreen assay, Alphascreen phosphotyrosine acceptor beads followedby streptavidin donor beads were added under subdued light. The ELISAplates were read on a BMG Fluorostar, the Alphascreen plates were readon a Packard Fusion Alpha. Inhibitors were added to the assays fifteenminutes prior to the addition of ATP. Inhibitors were added in aqueousDMSO, with DMSO concentrations never exceeding 1%.

Establishment of TEL:JAK Cell Lines

The coding region encompassing nucleotides 1-487 of TEL was amplified byPCR using the oligonucleotides 5TEL (5′-GGA GGA TCC TGA TCT CTC TCG CTGTGA GAC-3′) (SEQ ID NO:10) and 3TEL (5′-AGGC GTC GAC TTC TTC TTC ATG GTTCTG-3′) (SEQ ID NO: 11) and U937 mRNA as template. A BamH I site waspresent into the STEL Primer, a Sal I site was incorporated into the3TEL primer. The regions encompassing the kinase domains of JAK2(nucleotides 2994-3914; JAK2F 5′-ACGC GTC GAC GGT GCC TTT GAA GAC CGGGAT-3′(SEQ ID NO:12); JAK2R 5′-ATA GTT TAG CGG CCG CTC AGA ATG AAG GTCATT T-3′) (SEQ ID NO:13) and JAK3 (nucleotides 2520-3469; JAK3F 5′-GAAGTC GAC TAT GCC TGC CAA GAC CCC ACG ATC TT-3′ (SEQ ID NO:14); JAK3R5′-GGA TCT AGA CTA TGA AAA GGA CAG GGA GTG GTG TTT -3′) (SEQ ID NO: 15)were generated by PCR using Taq DNA Polymerase (Gibco/BRL) and U937 mRNAas template. A Sal1 site was incorporated into the forward primer ofJAK2 and JAK3, a Not I site was incorporated into the JAK2 reverseprimer and a Xba I site was added to the reverse primer of JAK3.

A TEL/Jak2 fusion was generated by digestion of the TELPCR product withBamH I/Sal I, digestion of the JAK2 PCR product with Sal I/Not Ifollowed by ligation and subcloning into the mammalian expression VectorpTRE 2 (Clontech) digested with BamH I-Not I (pTELJAK2). For JAK3 SalI/Not I cleaved kinase domain PCR product was ligated with BamH I/Sal Icleaved TELproduct followed by ligation into BamH I/Not I cleaved pTRE2(pTELJAK3).

The growth factor dependent myelomonocytic cell line BaF3 bearing thepTET-off plasmid (Clontech) was transfected with either pTELJAK2 orpTELJAK3 and the cells selected for factor independent growth. BaF 3wild type cells were cultured in DMEM 10% FCS, 10% WEHI 3B conditionedmedium. BaF3 TELJAK cells were cultured in DMEM 10% Tet-System ApprovedFBS (without WEHI 3B conditioned medium).

Cellular Assays were Performed as Follows:

Cell suspensions were prepared by harvesting cells from culture. (Cellsused in this test should be in later log phase growth and highviability.) Cells were diluted in correct growth medium to 1.1× finalconcentration (from 50000 cell/mL to 200,000 cell/mL, depending on cellline).

Compounds to be tested were added (10 μL, 10× final concentration) to aflat bottom 96-well plate. The cellular suspension (90 μL per well) wasadded, and the plate incubated for 40 hr at 37° C., 5% CO₂. MTT (20 μLper well, 5 mg/mL in PBS) was added and the palates were returned to theincubator for a further 6 hours. Lysis buffer (100 μL per well, 10% SDS,0.01N HCl) was added and the plate stored in the incubator overnight.The plate was then read at 590 nm.

Results

The activity of a range of compounds is shown in Table 3. Compounds thatexhibited a capacity to inhibit 50% of JAK activity at a concentrationof 50 μM (measured under standard conditions, see Methods), aredesignated as “+”.

It will be appreciated by persons skilled in the art that numerousvariations and/or modifications may be made to the invention as shown inthe specific embodiments without departing from the spirit or scope ofthe invention as broadly described. The present embodiments are,therefore, to be considered in all respects as illustrative and notrestrictive.

TABLE 3 Disubstituted pyrazines possessing JAK inhibitory activityCHEMISTRY Jak2 Jak3 abl ack kdr

+ − + + +

− − + + −

− + − + −

− − + + −

+ + + + −

+ − + + −

− − − − −

− + − + −

− − + NT +

− − + NT +

+ − + + +

+ − + + +

− − + + +

− − + + +

+ − + + −

+ − + + +

− − + + +

+ − − − −

− − − + −

− − + + −

− − + + +

− − + + +

+ − + + +

+ + + + +

+ − + + +

+ − + + +

− − − + +

− + − + −

− − + + −

+ − + + +

− − − − −

+ − + + +

− − − + −

− − + + +

− − + + +

+ − + − +

− − + + +

+ + + + +

+ − + + +

+ + + + +

+ − + + +

+ − + + +

− − + + +

− − + + −

+ − + + +

+ + + + +

+ + + + +

− − + + +

+ − + + +

+ − + + +

+ − + + +

− − + + +

− − + + +

− − + + +

+ + + + +

+ + + + +

+ − − + +

− − − + +

+ − − + +

+ + − + +

+ − + + +

+ − + + +

+ + + + +

+ + − + +

+ + + + +

+ − + + +

+ + + + +

+ + + + +

+ − + + +

+ + − − +

+ − + + +

+ + + + +

+ − + + +

+ + + + +

+ + − − +

+ + + + +

− − − + +

+ − + + +

+ + + + +

+ − − + +

− − + + −

+ − − + +

1. A compound of the formula:

or pharmaceutically acceptable salts, crystal forms or diastereomersthereof, wherein: D is:

where X₁, X₂, X₃, X₄ are optionally substituted carbon, or one of X₁,X₂, X₃, X₄ is N; R² is 0-4 substituents independently chosen from H,halogen, C₁₋₄ alkyl, CH₂F, CHF₂, CF₃, OCF₃, aryl, hetaryl, C₁₋₄alkylOC₁₋₄ alkyl, C₁₋₄ alkylOaryl, C₁₋₄ alkylNR³R⁴, CO₂R³, CONR³R⁴,CONR³SO₂R⁴, NR³R⁴, C₁₋₄ alkylNR³R⁴, nitro, NR³COR⁴, NR⁵CONR³R⁴,NR³SO₂R⁴, C₁₋₄ alkylNR³COR⁴, C₁₋₄ alkylNR⁵CONR³R⁴, or C₁₋₄alkylNR³SO₂R⁴; R³ and R⁴ are each independently H, halogen, CH₂F, CHF₂,CF₃, C₁₋₄ alkyl, C₁₋₄ alkyl cycloalkyl, C₁₋₄ cyclohetalkyl, aryl, C₁₋₄alkyl aryl, hetaryl, C₁₋₄ alkyl hetaryl, or may be joined to form anoptionally substituted 3-8 membered (saturated or unsaturated) ringoptionally containing an atom selected from O, S, NR⁶; R⁵ is H, C₁₋₄alkyl, halogen, CH₂F, CHF₂, CF₃, aryl or hetaryl; R⁶ is H, C₁₋₄ alkyl,aryl, hetaryl, C₁₋₄ alkyl aryl, or C₁₋₄ alkyl hetaryl; R¹ is H, C₁₋₄alkyl, C₁₋₆ cycloalkyl; Q is a bond, CH₂, C₁₋₄ alkyl; A is aryl, hetaryloptionally substituted with 0-3 substituents independently chosen fromhalogen, C₁₋₄ alkyl, CH₂F, CHF₂, CF₃, OCF₃, CN, NR⁸R⁹, aryl, hetaryl,C₁₋₄ aryl, C₁₋₄ hetaryl, C₁₋₄ alkylNR⁸R⁹, OC₁₋₄ alkylNR⁸R⁹, nitro,NR¹⁰C₁₋₄NR⁸R⁹, NR⁸COR⁹, NR¹⁰CONR⁸R⁹, NR⁸SO₂R⁹, CONR⁸R⁹, or CO₂R⁸, whereR⁸ and R⁹ are each independently H, C₁₋₄ alkyl, aryl or which togetherform an optionally substituted 4-8 membered ring which may contain aheteroatom selected from O, S, NR¹¹, where R¹¹ is C₁₋₄ alkyl, and R¹⁰ isselected from H, C₁₋₄ alkyl; and W is H, C₁₋₄ alkyl, or C₂₋₆ alkenyl,where C₁₋₄ alkyl or C₂₋₆ alkenyl may be optionally substituted with C₁₋₄alkyl, OH, OC₁₋₄ alkyl, NR¹²R¹³; and R¹², and R¹³ are each independentlyH, C₁₋₄ alkyl, or may be joined to form an optionally substituted 3-8membered ring optionally containing an atom selected from O, S, and NR¹⁴where R¹⁴ is H or C₁₋₄ alkyl.
 2. The compound of claim 1 of the formulaII

or pharmaceutically acceptable salts, crystal forms or diastereomersthereof, wherein: D is:

where X₁, X₂, X₃, X₄ are optionally substituted carbon, or one of X₁,X₂, X₃, X₄ is N; R² is 0-4 substituents independently chosen from H,halogen, C₁₋₄ alkyl, CH₂F, CHF₂, CF₃, OCF₃, aryl, hetaryl, C₁₋₄alkylOC₁₋₄ alkyl, C₁₋₄ alkylOaryl, C₁₋₄ alkylNR³R⁴, CO₂R³, CONR³R⁴,CONR³SO₂R⁴, nitro, NR³R⁴, C₁₋₄ alkylNR³R⁴, NR³COR⁴, NR⁵CONR³R⁴,NR³SO₂R⁴, C₁₋₄ alkylNR³COR⁴, C₁₋₄ alkylNR⁵CONR³R⁴, or C₁₋₄alkylNR³SO₂R⁴; R³ and R⁴ are each independently H, halogen, CH₂F, CHF₂,CF₃, C₁₋₄ alkyl, C₁₋₄ alkyl cycloalkyl, C₁₋₄ cyclohetalkyl, aryl, C₁₋₄alkyl aryl, hetaryl, C₁₋₄ alkyl hetaryl, or may be joined to form anoptionally substituted 3-8 membered (saturated or unsaturated) ringoptionally containing an atom selected from O, S, NR⁶; R⁵ is H, C₁₋₄alkyl, halogen, CH₂F, CHF₂, CF₃, aryl or hetaryl; R⁶ is H, C₁₋₄ alkyl,aryl, hetaryl, C₁₋₄ alkyl aryl, or C₁₋₄ alkyl hetaryl; R¹ is H, C₁₋₄alkyl, or C₁₋₆ cycloalkyl; W is H or C₁₋₄ alkyl; and A is aryl, hetaryloptionally substituted with 0-3 substituents independently chosen fromhalogen, C₁₋₄ alkyl, CH₂F, CHF₂, CF₃, OCF₃, CN, nitro, NR⁸R⁹, aryl,hetaryl, C₁₋₄ aryl, C₁₋₄ hetaryl, C₁₋₄ alkylNR⁸R⁹, OC₁₋₄ alkylNR⁸R⁹,NR¹⁰C₁₋₄NR⁸R⁹, NR⁸COR⁹, NR¹⁰CONR⁸R⁹, NR⁸SO₂R⁹, CONR⁸R⁹, or CO₂R⁸, whereR⁸ and R⁹ are each independently H, C₁₋₄ alkyl, aryl or which togetherform an optionally substituted 4-8 membered ring which may contain aheteroatom selected from O, S, and NR¹¹, where R¹¹ is C₁₋₄ alkyl, andR¹⁰ is H or C₁₋₄ alkyl.
 3. The compound of claim 1, where W is C₁₋₄alkyl wherein the compound possesses S chirality at the chiral carbonbearing W.
 4. The compound of claim 3, wherein the compound is a mixtureof R and S isomers and the mixture comprises at least 70% of the Sisomer.
 5. The compound of claim 4, wherein the compound comprises atleast 80% of the S isomer.
 6. The compound of claim 5, wherein thecompound comprises at least 90% of the S isomer.
 7. The compound ofclaim 6, wherein the compound comprises at least 95% of the S isomer. 8.The compound of claim 7, wherein the compound comprises at least 99% ofthe S isomer.
 9. The compound of claim 1, wherein the compound isselected from the group consisting of:N-benzyl-6-(1H-imidazol-1-yl)pyrazin-2-amine,N-(4-Fluorobenzyl)-6-(1H-imidazol-1-yl)pyrazin-2-amine,6-(1H-Imidazol-1-yl)-N-[(1R)-1-phenylethyl]pyrazin-2-amine,6-(1H-imidazol-1-yl)-N-(4-morpholin-4-ylphenyl)pyrazin-2-amine,N-[(1R)-1-Phenylethyl]-6-(4-phenyl-1H-imidazol-1-yl)pyrazin-2-amine, and6-(1H-Imidazol-1-yl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine.
 10. Thecompound of claim 1, wherein the compound is selected from the groupconsisting of:


11. A pharmaceutical composition comprising a pharmaceuticallyacceptable carrier and at least one compound of claim 1.